Antagonistic Isolates Research Articles

Plant, bacteria and fungi crosstalk: Direct and indirect biocontrol mechanisms of sugarcane rhizoplane Pseudomonas species against Fusarium wilt

In order to identify an effective rhizoplane Pseudomonas species to combat sugarcane wilt pathogen (Fusarium sacchari), twenty Pseudomonas isolates were obtained from the sugarcane rhizoplane of different genotypes. The three-tier confrontation (direct, indirect and remote) tests revealed antagonistic activity of the bacteria, bacterial non-volatile and volatile compounds. In dual culture assay, SRP19 (46.30%) and SRP20 (39.30%) have shown the highest inhibition against F. sacchari. 16S-23S rDNA sequencing identified them as Pseudomonas chlororaphis subsp. aurantiaca (SRP19) and Pseudomonas aeruginosa (SRP20). Exploration of biocontrol mechanisms revealed the potential roles for IAA production, extracellular enzyme secretion (proteases, cellulases and chitinases), HCN synthesis, siderophore production, pyocyanin or fluorescein production and phosphate solubilization. The molecular analysis revealed the presence of genes (phz, prnD, phlD and pltC) involved in the biosynthesis of antimicrobial peptides like phenazines, pyrrolnitrin, 2,4-diacetyl phloroglucinol and pyoleuteorin, involved in suppressing Fusarium. The antagonistic activity was further confirmed under greenhouse conditions by sett treatment with potential antagonistic isolates, which increased germination and reduced the pre-emergent mortality by 60% and post-emergent seedling mortality by 76% in SRP19 treated pots. The study revealed that the isolate Pseudomonas chlororaphis subsp. aurantiaca strain SRP19 can act as PGPR with both plant growth promoting and pathogen suppressing activity that can be used as an effective biocontrol agent against Fusarium wilt of sugarcane.

Sustainable management of peanut damping-off and root rot diseases caused by Rhizoctonia solani using environmentally friendly bio-formulations prepared from batch fermentation broth of chitinase-producing Streptomyces cellulosae.

Soil-borne plant diseases represent a severe problem that negatively impacts the production of food crops. Actinobacteria play a vital role in biocontrolling soil-borne fungi. The target of the present study is to test the antagonistic activity of chitinase-producing Streptomyces cellulosae Actino 48 (accession number, MT573878) against Rhizoctonia solani. Subsequently, maximization of Actino 48 production using different fermentation processes in a stirred tank bioreactor. Finally, preparation of bio-friendly formulations prepared from the culture broth of Actino 48 using talc powder (TP) and bentonite in a natural as well as nano forms as carriers. Meanwhile, investigating their activities in reducing the damping-off and root rot diseases of peanut plants, infected by R. solani under greenhouse conditions. Actino 48 was found to be the most significant antagonistic isolate strain at p ≤ 0.05 and showed the highest inhibition percentage of fungal mycelium growth, which reached 97%. The results of scanning electron microscope (SEM) images analysis showed a large reduction in R. solani mycelia mass. Additionally, many aberrations changes and fungal hypha damages were found. Batch fermentation No. 2, which was performed using agitation speed of 200rpm, achieved high chitinase activity of 0.1163 U mL- 1 min- 1 with a yield coefficient of 0.004 U mL- 1 min- 1 chitinase activity/g chitin. Nano-talc formulation of Actino 48 had more a significant effect compared to the other formulations in reducing percentages of damping-off and root rot diseases that equal to 19.05% and 4.76% with reduction percentages of 60% and 80%, respectively. The healthy survival percentage of peanut plants recorded 76.19%. Furthermore, the nano-talc formulation of Actino 48 was sufficient in increasing the dry weight of the peanut plants shoot, root systems, and the total number of peanut pods with increasing percentages of 47.62%, 55.62%, and 38.07%, respectively. The bio-friendly formulations of actinobacteria resulting from this investigation may play an active role in managing soil-borne diseases.

Isolation of Potential Antagonistic Rhizosphere Fungi against Alternaria alternata from Organic Carrot Productions

Rhizospheric fungi can have serve as biological agents against plant pathogens Alternaria alternata, which infects carrot plants. Rhizospheric fungi are more abundant in organic lands. This study aims to isolate fungi from organic carrot’s rhizosphere that are able to suppress A. alternata’s growth and determine isolate rhizospheric fungi ability to inhibit A. alternata to identify potential biological control agents against A. alternata. This research was conducted at the Laboratory of Biotechnology and Basic Biology, Diponegoro University, between January–June 2023. Research methods include survey location, soil sampling, rhizospheric fungi isolation, pathogenic fungi isolation, creating growth curve, pathogenicity tests, antagonism tests, and fungi identification. This study used a complete randomized design (CRD) and was analyzed using ANOVA at the 5% level of significance and Duncan's hoc-posttest. Fungi isolation from organic carrot plant’s rhizosphere resulted in 13 isolates consisting of six genera: Penicillium, Aspergillus, Paecilomyces, Myrothecium, Trichoderma, and Simplicillium. The antagonism test showed that eight antagonistic isolates, including the genus Penicillium (WO1, WO6, WO10), Paecilomyces (WO5), Myrothecium (WO7), Trichoderma (WO9), and Aspergillus (WO11, WO13), where WO1, WO6, WO10, and WO11 have a medium percentage of inhibition (44.12-57.84%), while WO5, WO7, WO9, and WO13 have a high percentage of inhibition (64.29-76.06%). The eight isolates showed antagonism mechanisms, including antibiosis (WO1, WO5, WO6, WO7, WO10, and WO11) and parasitism (WO9 and WO13).

Biocontrol potential of epiphytic bacteria against Xanthomonas citri pathotypes A and A*

BackgroundCitrus canker caused by the bacterium Xanthomonas citri (Hasse 1915) is known as a dangerous disease that has serious economic impacts on citrus production in many tropical and subtropical countries. In Iran, Citrus canker disease has caused significant damage to various citrus fruits, particularly limes. Nowadays, employing antagonistic bacteria as an effective and environmentally friendly method for plant disease management has gained special importance.ResultsCitrus leaf samples were collected from different regions of Iran, and after isolating epiphytic bacteria, ex planta and in planta investigations concerning antagonistic activities of the isolates against Xanthomonas citri pathotypes A and A* were performed. A total of 94 epiphytic bacterial isolates were isolated from citrus aerial parts. Based on biochemical, physiological, morphological, and genotypic tests, it was determined that these bacteria belong to the Bacillus and Staphylococcus genera. The highest inhibition activity against the pathogenic bacterium was related to isolates D4 and D5. Using the molecular method and the resulting dendrogram, it was found that these isolates were most similar to Bacillus amyloliquefaciens. The present findings demonstrated that pathogenicity test on key lime leaves infected with X. citri pathotypes A and A*, along with the application of the biocontrol strain B. amyloliquefaciens, resulted in a significant reduction in the number of canker lesions.ConclusionThe results strongly suggested that the identified antagonistic bacterial isolates hold promising potential as biocontrol agents for managing citrus bacterial canker disease.

Antifungal potential of volatiles produced by Aspergillus neoniger F14 against Ganoderma boninense

Ganoderma boninense Pat. is a fungal pathogen that primarily afflicts oil palm, leading to decreased palm oil yield and significant economic losses. The recent focus on biocontrol agents producing acids and volatile compounds has gained momentum due to their straightforward synthesis and ease of interaction with the phytopathogen. This research is a component of a project aimed at exploring the local biodiversity of antagonistic fungi in the oil palm plantation environment, with a specific focus on volatile organic compounds (VOCs) detected using Gas Chromatography-Mass Spectrometry (GC-MS). Isolate F14, a collection in the Laboratory of Applied Microbiology, BRIN, Indonesia was identified as Aspergillus neoniger, a novel report as a soil-originating fungal community in Indonesian oil palm plantations. VOC profiling using GC-MS revealed five dominant compounds namely (Z)-9-Tricosene, 1,2,3,4,5-Pentamethylcyclopentane, Dihexyl carbonate, Pentadecane, and Tridecyl laurate. Based on aromatogram disc tests against G. boninense colonies, it was determined that VOCs from A. neoniger F14 inhibited growth by 48% on the fifth day. The findings of this study contribute supportive information for further investigations into VOCs produced by other local antagonistic isolates against G. boninense.

Field performance assessment of formulated Pseudomonas fluorescens for enhancing plant growth and inducing resistance against rice blast disease

To control blast disease in the rice field under natural conditions, three antagonistic isolates of rhizospheric P. fluorescens were formulated in talc, kaolinite, PVP, and vegetable oil using an RCBD with three replications. At various phases of the rice plants' growth, all of the products markedly accelerated plant growth and yield-contributing characteristics. Pf-8, compounded in talc (5 %) and vegetable oil (2 %) among the three isolates of P. fluorescens, significantly elevated vegetative and yield parameters with higher (2.11 and 2.08) benefit-cost ratios, respectively. At 90 days after transplanting (DAT), T11 (5 % Pf-8 Talc) and T13 (2 % Pf-8 Vegetable oil) showed a significant reduction in blast incidence (76.81 %, 75.45 %) and severity (71.57 %, 69.82 %), with the largest populations of P. fluorescens (9.60 × 1010 and 9.51 × 1010) in the rhizosphere. Moreover, a significantly increased level of phenol and hydrogen peroxide (H2O2) was found in Pseudomonas-treated leaves (Pf-8) at 30, 60, and 90 DAT indicating a strong relationship with rice blast disease reduction. Further, blast incidence and severity showed a negative correlation with vegetative parameters, yield parameters, phenol, and H2O2. Thus, it may be claimed that using formulated P. fluorescens (Pf-8) to manage rice blast in the field could be an alternate strategy to using chemicals.

Internal Transcribed Spacers (ITS) Based Identification of Trichoderma Isolates and Biocontrol Activity against Fusarium oxysporum and Chitinase Activity

Twelve Trichoderma spp. isolated from different locations in Uttar Pradesh, India. Internal transcribed spacer (ITS) amplification and sequencing were used to identify isolates of Trichoderma that showed 99–100% identification with Trichoderma harzianum, Trichoderma asperellum, and Trichoderma longibrachiatum, the three species of Trichoderma. Tests were conducted in Vitro to evaluate the biocontrol potential of Trichoderma isolates against Fusarium species. After seven days of incubation, the isolate TBT6 (T. harzianum) exhibited the greatest antagonism against Fusarium oxysporum, with inhibition rates of 87.1%) by TBT7 (82.2%), and the least amount of inhibition (59.7%) by T. longibrachiatum isolate TBT10. Under field conditions, the antagonistic T. harzianum isolate TBT6 can be employed as a bio-control agent against Fusarium oxysporum. and for the development of formulations based on Trichoderma. Using the dual plate approach, twelve highly antagonistic Trichoderma isolates were selected for chitinolytic activity. Observing the breakdown of chitin substrates, Trichoderma isolates TBT6 was shown to have strong chitinolytic enzyme activity, making it a good candidate for endochitinase gene isolation. Using two particular primers, the genomic DNA of the Trichoderma isolate TBT6 was isolated and amplified.

Evaulation of Effective Antagonistic Yeasts in Kalasin Province, a Biocontrol Agent against the Fungal Colletotrichum capsici Caused Anthracnose Diseases in Chili Fruits

Primary screening of epiphytic and endophytic yeasts, the majority yeast was found from rhizosphere soil (21 isolates, 31.81%), and 5 antagonistic yeasts can have inhibited the growth of mycelia of C. capsici up to 60%. Efficacy of five antagonistic yeasts were screened under plant nurseries conditions. The result found all isolates were tested and compared with control and chemical plots. The result of plant height showed that, yeast isolate KS34 had highest of plant height at 45.80 cm. For the result of branch per plant of chili plant found isolate KS37 showed highest of branch per plant at 16.60 branches and the flower of chili per plant presented that isolate KS37 had highest of flower at 18.60 flowers. The result of anthracnose disease control by antagonistic yeasts in disease incidence and category mean as plant resistance. These results found that, Isolates KS46 had a great disease incidence at 7.80% that mean this isolate prove chili plant to resistance to C. capsisci. Addition, isolate KS46 had highest of activity of chitinase enzyme at 170.72 U/ml. Isolate KS46 identified by D1/D2 domain of 26S ribosomal DNA sequencing. The sequence analysis showed maximum identity of 100% with Candida haemulonii. This assumed C. haemulonii was showed the best to control C. capsici. For confirmed this result, all isolates were determined the mode of action by studied in relationship between antagonist isolates. The action of yeast isolate could competition the nutrient with mycelia of pathogen. This mode effect to C. capsici loss nutrient caused the pathogen get stuck to braked and died later.

Production, optimization and characterization of partially purified anti-mycotic compound from marine soil derived streptomycetes originating at unexplored region of Bay of Bengal, India

Sixty-eight morphologically distinct isolates of marine actinomycetes were derived from seashore, mangrove, and saltpan ecosystems located between the Palk Strait and Gulf of Mannar region, Bay of Bengal, Tamilnadu. Twenty-five (36.8%) isolates exhibited anti-mycotic activity against Candida albicans and Cryptococcus neoformans in preliminary screening, and 4 isolates with prominent activity were identified and designated at the genus level as Streptomyces sp. VPTS3-I, Streptomyces sp. VPTS3-2, Streptomyces sp. VPTSA1-4 and Streptomyces sp. VPTSA1-8. All the potential antagonistic isolates were further characterized with phenotypic and genotypic properties including 16S rRNA gene sequencing and identified species level as Streptomyces afghaniensis VPTS3-1, S. matensis VPTS3-2, S. tuirus VPTSA1-4 and S. griseus VPTSA1-8. In addition, the active fractions from the potential antagonistic streptomycetes were extracted with organic solvents by shake flask culture method and the anti-mycotic efficacies were evaluated. The optimization parameters for the production of the anti-mycotic compound were found to be pH between 7 and 8, the temperature at 30ᵒC, the salinity of 2%, incubation of 9 days, and starch and KNO3 as the suitable carbon and nitrogen sources respectively in starch casein medium.

Exploring Fungal Candidates from Aromatic Rhizosphere as Biological Controls Against Three Pyricularia oryzae Haplotypes.

<b>Background and Objective:</b> Blast disease (<i>Pyricularia oryzae</i>) is a major disease-causing yield losses in rice crops worldwide. Disease control using resistant varieties is less effective due to the high genetic variation in <i>P. oryzae</i> populations in the field and the use of synthetic fungicides hurts the diversity of biological agents. This study aims to explore fungi in the rhizosphere of organic aromatic rice in North Luwu Regency that can utilized as biological control agents against three haplotypes of <i>P. oryzae</i>. <b>Materials and Methods:</b> Isolation of rhizosphere fungi using serial dilution method and scatter plate method. The identification of fungi based on microscopic and macroscopic characteristics. Genotype test of 15 <i>P. oryzae</i> isolates used gene-based markers related to virulence traits, namely Erg2 (1,440 bp), Pwl2 (900 bp) and Cut1 (1,730 bp). Amplified DNA bands that appeared were scored as 1 (present) and 0 (absent). <b>Results:</b> Exploring organic rice rhizosphere fungi in North Luwu Regency found potential biological control agents against three <i>P. oryzae</i> haplotypes on local varieties: Juvenile and Bandarata. Twelve fungal isolates from the rhizosphere of aromatic rice were successfully isolated and six antagonistic fungal isolates were able to inhibit the growth of <i>P. oryzae</i> haplotypes C-011, D-111 and F-110. <i>Trichoderma</i> spp., isolates had the highest inhibition percentage of 72-90%, followed by <i>Penicillium </i>sp., 1 with an inhibition percentage of 62-82%. <b>Conclusion:</b> Twelve fungal isolates from the rhizosphere of aromatic rice were successfully isolated and six antagonistic fungal isolates were able to inhibit the growth of <i>P. oryzae</i> haplotypes C-011, D-111 and F-110.

Evaluation of phyllosphere bacterial biocontrol agents for the suppression of rice foliar diseases

AbstractPhyllosphere colonizing antagonistic bacterial isolates were evaluated for managing major rice foliar diseases, namely brown spot caused by Bipolaris oryzae and sheath rot caused by Sarocladium oryzae. Among these various isolates, Pseudomonas aeruginosa isolate 1 effectively inhibited the mycelial growth of B. oryzae and S. oryzae. The volatile compounds of Bacillus subtilis isolates effectively inhibited the mycelial growth of S. oryzae and B. oryzae. Non‐volatile compounds of P. aeruginosa isolate 2 and B. subtilis isolate 1 recorded the maximum inhibition of mycelial growth of B. oryzae and S. oryzae, respectively. P. aeruginosa isolates produced fluorescent pigments and hydrogen cyanide (HCN), whereas isolates of Bacillus spp. did not. All the isolates of P. aeruginosa and Bacillus spp. produced protease enzymes. P. aeruginosa isolates solubilized phosphate, whereas isolates of Bacillus spp. did not. Rice seeds treated with these bacterial suspensions showed improvement in plant growth parameters. The foliar application of two bacterial strains, namely P. aeruginosa isolate 1 + B. subtilis isolate 1 recorded the minimum brown spot and sheath rot disease incidence. Thus, the present showed that phyllosphere colonizing P. aeruginosa and B. subtilis have the potential application for managing foliar fungal diseases of rice plants.

Production of Pumilarin and a Novel Circular Bacteriocin, Altitudin A, by Bacillus altitudinis ECC22, a Soil-Derived Bacteriocin Producer.

The rise of antimicrobial resistance poses a significant global health threat, necessitating urgent efforts to identify novel antimicrobial agents. In this study, we undertook a thorough screening of soil-derived bacterial isolates to identify candidates showing antimicrobial activity against Gram-positive bacteria. A highly active antagonistic isolate was initially identified as Bacillus altitudinis ECC22, being further subjected to whole genome sequencing. A bioinformatic analysis of the B. altitudinis ECC22 genome revealed the presence of two gene clusters responsible for synthesizing two circular bacteriocins: pumilarin and a novel circular bacteriocin named altitudin A, alongside a closticin 574-like bacteriocin (CLB) structural gene. The synthesis and antimicrobial activity of the bacteriocins, pumilarin and altitudin A, were evaluated and validated using an in vitro cell-free protein synthesis (IV-CFPS) protocol coupled to a split-intein-mediated ligation procedure, as well as through their in vivo production by recombinant E. coli cells. However, the IV-CFPS of CLB showed no antimicrobial activity against the bacterial indicators tested. The purification of the bacteriocins produced by B. altitudinis ECC22, and their evaluation by MALDI-TOF MS analysis and LC-MS/MS-derived targeted proteomics identification combined with massive peptide analysis, confirmed the production and circular conformation of pumilarin and altitudin A. Both bacteriocins exhibited a spectrum of activity primarily directed against other Bacillus spp. strains. Structural three-dimensional predictions revealed that pumilarin and altitudin A may adopt a circular conformation with five- and four-α-helices, respectively.

Population dynamics of Trichoderma harzianum in bio- fortified compost against soil-borne potato diseases

Soil-borne diseases pose significant challenges to global potato cultivation, resulting in yield losses and economic implications. Utilizing biological control agents offers a sustainable and environmentally and friendly approach to manage these diseases. Before implementing the experiment in the field, a series of in-vitro tests were conducted to select a virulent isolate of the tested pathogens and an effective antagonistic isolate of Trichoderma harzianum. In the pathogenicity test, isolate RS-1 of Rhizoctonia solani and isolate SR-8 of Sclerotium rolfsii exhibited the highest virulence against two pathogens in potato variety ‘Cardinal’ in regard to seedling mortality (44.44-88.88%). During in vitro screening, T. harzianum isolate Tri 7 displayed the greatest ability to inhibit the mycelium growth of all tested pathogens. In the field trial, the highest seedling mortality (69.06%) was observed in T2 (Fresh seeds+ Soil inoculated with R. solani and S. rolfsii). However, the highest seedling mortality (86.94%) was reduced by treatment T7 (Fresh seeds+ Soil inoculated with R. solani and S. rolfsii +500g wheat grain colonized with Trichoderma fortified poultry compost). Treatment T7 also recorded the lowest incidences of stem canker (7.83%) and stem rot (8.64%), whereas the highest disease incidences occurred in the T2 treatment plot. Notably, treatment T7 gave the highest yield of 30.94 tons/ha, while the control plot T2 yielded the lowest at 12.17 tons/ha. Initially, Trichoderma population was minimal prior to the application of Trichoderma- fortified compost. However, populations gradually increased over time, peaking at 3 months across all treatments except the control, where no Trichoderma was applied. However, a decline in T. harzianum population was noted one month after potato harvesting. This approach offers a potential solution for managing potato soil-borne diseases in a sustainable and environmentally friendly manner. Ann. Bangladesh Agric. (2023) 27 (1): 81-92

Evaluation of antifungal activity against Corynespora cassiicola by bacteria isolated from soil in the root zone of cucumber plants

Abstract. Duy LQ. 2023. Evaluation of antifungal activity against Corynespora cassiicola by bacteria isolated from soil in the root zone of cucumber plants. Biodiversitas 24: 6584-6591. Target leaf spot caused by Corynespora cassiicola poses a major disease threat to cucumber cultivation worldwide. Reliance on chemical fungicides has raised resistance to environmental and health concerns. As cucumber production expands globally, sustainable solutions are needed. Plant Growth-Promoting Rhizobacteria (PGPR) show promise but remain underexplored against C. cassiicola in cucumber. This study aimed to assess the potency of rhizobacteria in antagonistic activity against C. cassiicola, causing cucumber target leaf spots under in vitro conditions. The present study isolated bacterial strains from cucumber soil rhizosphere and screened for antifungal activity against C. cassiicola using dual culture assays. Using quantitative assays, selected antagonistic strains were further assessed for key biocontrol traits, including ?-1,3-glucanase, chitinase, and siderophore production. Cell-free supernatants were tested for direct fungal growth inhibition using the environmental toxicity. Isolated PGPR strains showed robust C. cassiicola inhibition from 39.8% to 62.6% growth reduction. Most antagonistic isolates produced ?-1,3-glucanase, chitinase, and siderophores and suppressed pathogen growth via metabolites in their supernatants. These results demonstrate multiple biocontrol mechanisms, including enzymatic degradation of fungal cell walls, iron competition through siderophores, and antimicrobial compound secretion. Bacillus siamensis strain TV16 exhibited the strongest overall antagonism against C. cassiicola. These findings reveal the biocontrol potential of PGPR for managing cucumber-target leaf-spot disease, offering a sustainable alternative to chemical pesticides. With further optimization and field testing, PGPR-based biopesticides can be integrated into cucumber production to improve crop health while reducing fungicide reliance.

Efficacy of Seed-Biopriming with Trichoderma spp. and Foliar Spraying of ZnO-Nanoparticles Induce Cherry Tomato Growth and Resistance to Fusarium Wilt Disease.

Several microbes that cause plant diseases drastically lower the production of agriculture and jeopardize the safety of the world's food supply. As a result, sustainable agriculture requires disease management tactics based on modern, eco-friendly techniques as alternatives to various agrochemicals. The current study aimed to assess the antifungal activity of ZnO-nanoparticles against Fusarium solani in-vitro, and the ability of two antagonistic Trichoderma isolates, Trichoderma viride and Trichoderma harzianum, to produce antifungal secondary metabolites and identify them using gas chromatography-mass spectrometry, and to evaluate the combined effects of foliar spray of ZnO-nanoparticles and bioprimed seeds of cherry tomato (Solanum lycopersicum L.) with two antagonistic Trichoderma isolates against Fusarium wilt disease caused by Fusarium solani in greenhouse conditions. The results revealed that, in-vitro, the highest concentration of ZnO nanoparticles (3000 ppm) resulted in the greatest decrease in Fusarium solani mycelial growth (90.91% inhibition). The scanning electron microscopy demonstrated the evident distortion in Fusarium solani growing mycelia treated with ZnO-nanoparticles, which might be the source of growth suppression. Additionally, twenty-eight bioactive chemical compounds were isolated and identified from Trichoderma spp. ethyl acetate crude extracts using gas chromatography-mass spectrometry. In a greenhouse experiment, the combination of bioprimed cherry tomato plants with Trichoderma harzianum and foliar spraying of ZnO-nanoparticles at 3000 ppm was the most effective interaction treatment for reducing disease severity index (23.4%) and improving the vegetative growth parameters, micronutrient contents (Mn, Zn, and Fe in leaves), and chlorophyll content (SPAD unit), as well as stimulating phenylalanine ammonia-lyase activity of cherry tomato leaves at 75 days after sowing. In conclusion, the antifungal potential of seed-biopriming with antagonistic Trichoderma isolates and the foliar spraying of ZnO-nanoparticles can boost cherry tomato growth and confer resistance to Fusarium wilt caused by Fusarium solani.

Screening of Rhizobacterial Isolates from Apple Rhizosphere for Their Biocontrol and Plant Growth Promotion Activity

Apple crops are prone to several diseases that limit their production—in particular, root rot caused by a new genus of oomycetes, mainly Phytopythium vexans. This study aims to screen antagonistic bacteria that can play an important role in the biological control of this pathogenic oomycete and to evaluate their capacity to promote plant growth. The dual culture test revealed that, out of 200 bacterial isolates, 16 have been able to inhibit the mycelial growth of P. vexans with inhibition rates greater than 50%. The selected isolates were identified based on the 16S rDNA genes: 14 bacteria belonging to the genus Bacillus, Stenotrophomonas, and the family Enterobacteriaceae. Notably, two isolates, B1 and M2-6 (identified as Bacillus velezensis), demonstrated the highest inhibition rates of 70% and 68%, respectively. These selected isolates were examined for their ability to produce different compounds related to biocontrol and plant growth promotion. Furthermore, the 16 selected isolates were evaluated for their ability to produce compounds associated with biocontrol and plant growth promotion, including hydrolytic enzymes (cellulases, proteases, and amylases), HCN (hydrogen cyanide) production, phosphate solubilization, IAA (indole-3-acetic acid) production, pectinase production, and stimulation of sorghum bicolor growth in vivo. Variations were observed among the bacterial isolates in terms of their compound production and phytostimulation capabilities. However, the secretion of proteases was consistently detected in all antagonistic isolates. The presence of genes responsible for the production of antifungal lipopeptides (bacillomycin, fengycin, and iturin) in the selected bacterial isolates was determined using polymerase chain reaction (PCR) techniques, while the absence of genes involved in surfactin biosynthesis was also confirmed through PCR studies. These isolates demonstrated inhibitory activity through the production of proteases and antifungal lipopeptides. Further research is needed to explore their potential use in biological control strategies and to improve apple crop productivity.

Uji Antagonis Cendawan Isolat Lokal Terhadap Fusarium spp, Penyebab Penyakit Layu Pada Tanaman Kentang (Solanum tuberosum L ) Secara In Vitro.

This research aims to: (a) determine the number of fungal isolates capable of inhibiting the growth of Fusarium sp. in vitro, (b) determine the percentage of inhibitory activity of the antagonistic fungal isolates against Fusarium sp. in vitro, (c) identify the morphological characteristics of the top three antagonistic fungal isolates from the rhizosphere of potato plants (Solanum tuberosum L.). To achieve these objectives, the in vitro experimental testing was conducted at the Biology Laboratory of Makassar State University. The research results showed that: (a) based on macroscopic and microscopic morphological characteristics, the fungi isolated from potato plants were identified as Fusarium oxysporum f.sp. solanaceae. (b) There were seven isolates capable of inhibiting the growth of F.o.sp. solanaceae, namely Aspergillus sp (green), Aspergillus sp (black), Eupenicillium sp, Chaetomium sp, Paecilomyces sp, and Monilia sp. The percentage of their inhibitory activity was 54.71%, 73.13%, 36.82%, 55.01%, 50.66%, 46.53%, and 43%, respectively. (c) The morphological characteristics of the top three antagonistic fungal isolates were as follows: 1. Black Aspergillus sp had a black colony with a solid texture and very rapid growth. The hyphae were septate and non-branched, and the spherical conidia were black. 2. Green Aspergillus sp had a colony with dark green (inner), light green (middle), and white (outer) regions. The conidia were spherical, with a solid and slightly elevated texture, spreading in all directions. The hyphae were septate and non- branched. 3. Chaetomium sp had a white cotton-like colony with a fine and thick texture, spreading in all directions. The conidiophores were irregularly branched, and the hyphae were septate.

Test Fungi Growth Physiology of Trichoderma sp. and Gliocladium sp. from Citrus Plants

One of the problems of citrus cultivation is disease attack which can cause a decrease in production during harvest. There are antagonistic fungi from citrus plants that can be used to control pathogens, namely Gliocladium sp. and Trichoderma sp. Antagonistic fungi will be able to control pathogens optimally if they have growth conditions that match their environment. This study aims to determine the growth development of Trichoderma sp. and Gliocladium sp. at several temperatures, media, exposure lengths, and types of light. The treatments used were four different temperatures (20°C, 25°C, 30°C, 35°C), three media (PDA, PDAY, and CMA), several exposure times (24 hours of light and 24 hours of dark, 8 hours of light and 16 hours of darkness, 16 hours of light and 8 hours of darkness) and the type of light (light from a reading lamp placed very close, light from a lamp placed habitually, and dark wrapped in carbon). The treatment used a completely randomized design (CRD) and was repeated 5 times. The test results showed that the two isolates could develop optimally on PDA and PDAY media with the best temperature of 20°C-25°C. In addition, testing of various rays showed the results of the fastest development by Trichoderma sp. under natural light treatment. Furthermore, the best length exposure test was Trichoderma sp. with a lighting time of 12 hours light and 12 hours dark. In general, the development of the two antagonistic fungal isolates had relatively the same optimal development conditions, but Trichoderma sp. has better development on various tests.

Hemolysis and hypersensitive tests ease culture collection management of antagonistic bacteria

A biosafety assessment is a crucial first step in the management of microbial culture collection to screen and determine unexpected potential plant and human pathogenic bacteria. It is common to collect and store as many fresh culture collections from natural resources before being further evaluated for antagonist bacteria. As a result, a bulk of isolates must be preserved which required more effort and budget. Safety evaluations based on the hemolysis and hypersensitive reactions offer simple tests to ease culture collection management of antagonist bacteria. The study aimed to evaluate the safety of bacterial culture collection for their hemolysis and hypersensitive reactions. Ninety-five isolates of rhizosphere and endophytic bacterial isolates from the culture collections of the Department of Plant Protection-IPB University, Indonesian Industrial and Beverages Crops Research Institute, and Indonesian Spice and Medicinal Crops Research Institute, were evaluated their safety using the hemolysis and hypersensitive tests. The hemolysis test was conducted using blood agar media, from which isolates with a negative (?) reaction were then tested for the hypersensitivity reaction on tobacco leaves. Bacterial isolates passed from both hemolysis and hypersensitivity tests were then preserved by the lyophilization method for long-term storage of culture collection. Based on the hemolysis test, 68 out of 95 bacterial isolates (71.57%) were found to be positive (? or ?) reactions. The hypersensitive test showed that 22 of 27 negative hemolysis isolates did not trigger hypersensitivity reactions in tobacco leaves, therefore, they were preserved by lyophilization. The study indicated that a high number of bacterial isolates in the present collection, 68 positive hemolysis, and 5 hypersensitive, need to be re-evaluated due to their safety concerns. The present study highlights the importance of biosafety tests performed in an early stage before the to permanent collection of antagonist isolates.

Identification and characterization of Aspergillus niger causing collar rot of groundnut (Arachis hypogaea)

Abstract. Nguyen XH, Nguyen TMN, Nguyen DH, Nguyen QC, Cao TT, Pham TTH, Nguyen TTT. 2023. Identification and characterization of Aspergillus niger causing collar rot of groundnut (Arachis hypogaea). Biodiversitas 24: 2556-2562. Groundnut (Arachis hypogaea L.) is an economically important legume crop in Vietnam and many other countries worldwide. Groundnut cultivation is affected by many biotic and abiotic stresses. Among biotic stresses, groundnut is attacked by many fungal, bacterial and viral pathogens. The most harmful fungal diseases are rust, stem rot, collar rot and other soil borne diseases. Collar rot caused by Aspergillus niger is one of the most important disease of groundnut extensive in Vietnam and worldwide. In this study, symptomatic infected plant samples were collected from nine different locations in Quang Tri and Thua Thien Hue provinces, Vietnam Isolated fungal species were identified on the basis of morphological characterization, and molecular level. The pathogenicity of fungal species was confirmed on the basis of Koch's postulates. The result showed that a total of nine fungal strains were isolated from infected groundnut samples. Morphological analysis results showed that all isolates exhibited A. niger characteristics, including black colony, biseriate conidia, hyaline conidiophores and sterigmata. Pathogenicity test showed that all nine A. niger isolates caused collar rot disease in groundnuts. Further molecular identification based on ITS1-4 region nucleotides comparison indicated that selected two isolates, namely QT1 and H17 belonged to A. niger. Further research should be done to find antagonistic microorganisms isolates for the bio-control of A. niger.