An understanding of biocontrol activities is important when developing microorganism‐based alternatives to conventional fungicides. From our bacterial collection, we selected two strains (BBC023 and BBC047) for their outstanding antagonistic capacity against fungal phytopathogens and growth‐promoting abilities towards Arabidopsis thaliana. According to physiological and molecular characterizations, both strains were classified as Bacillus amyloliquefaciens and were tested against Botrytis cinerea in vitro and in a tomato. Both strains secrete lipopeptide‐like compounds that contribute to their in vitro antagonism. SEM‐images showed altered B. cinerea mycelial structures that were consistent with previous reports of the direct action of lipopeptides against fungal hyphae. The strains were applied to the roots (R), leaves (foliar ‐ F) or root/leaves (R/F) on tomato plants. All treatments significantly reduced the severity of B. cinerea infection (measured as a control index). However, only root applications (R and R/F) led to growth promotion in the tomato plants. We detected the production of indole acetic acid (IAA) and 2,3‐butanediol as growth promotion traits in the two strains. For both strains, the R/F treatment showed the highest control index, suggesting a synergic effect of direct antagonism against B. cinerea and resistance induction in the plant. In addition, in vitro antagonism of BBC023 and BBC047 against B. cinerea was similar; whereas in the F application, strain BBC047 significantly improved plant resistance and maintained a higher population density over time on tomato leaves, compared to BBC023. BBC047 was also able to produce a complex and robust biofilm in Msgg medium compared with that of BBC023. We linked the reduced biocontrol of BBC023 on leaves with its limited ability to generate robust biofilms and colonize the phylloplane. At last, we highlight the potential of the native Bacillus strains as promising alternatives for the development of bioproducts for sustainable agriculture.
Many aspects regarding the role of lipopeptides (LPs) in bacterial interaction with plants are not clear yet. Of particular interest is the LP family of surfactin, immunogenic molecules involved in induced systemic resistance (ISR) and the bacterial colonization of plant surfaces. We hypothesize that the concentration of surfactin produced by a strain correlates directly with its ability to colonize and persist on different plant surfaces, which conditions its capacity to trigger ISR. We used two Bacillus velezensis strains (BBC023 and BBC047), whose antagonistic potential in vitro is practically identical, but not on plant surfaces. The surfactin production of BBC047 is 1/3 higher than that of BBC023. Population density and SEM images revealed stable biofilms of BBC047 on leaves and roots, activating ISR on both plant surfaces. Despite its lower surfactin production, strain BBC023 assembled stable biofilms on roots and activated ISR. However, on leaves only isolated, unstructured populations were observed, which could not activate ISR. Thus, the ability of a strain to effectively colonize a plant surface is not only determined through its production of surfactin. Multiple aspects, such as environmental stressors or compensation mechanisms may influence the process. Finally, the importance of surfactin lies in its impacts on biofilm formation and stable colonization, which finally enables its activity as an elicitor of ISR.
BACKGROUND Consumption of bell peppers is recommended because of their bioactive compound content and their positive effects on health. Growth‐promoting rhizobacteria are popular because of their ability to promote plant growth by improving the fixation of nutrients or by inducing a systemic response. Green bell pepper (Capsicum annum) roots were inoculated with an autochthonous strain of Bacillus amyloliquefaciens, at different stages of development: T1, inoculation in the seedbed before transplant; T2, inoculation at and after transplant; T3, inoculation in the seedbed, at and after transplant. Bell pepper plants without inoculation were considered as control. Physicochemical composition and antioxidant activity of the fruits were measured to select the best treatment. RESULTS T1 increased crude proteins, fat, Ca, Fe, vitamin C, total phenolic content, antioxidant capacity by DPPH and by ORAC. On the other hand, T1 decreased reducing sugars, K and Cu content. No significant differences for total carbohydrates, ash and photosynthetic pigments were found. CONCLUSION Inoculated green bell peppers have enhanced its functional value and could be considered as an important source of bioactive compounds with elevated antioxidant activity. © 2019 Society of Chemical Industry
Tomato (Solanum lycopersicum L.) is an important crop in the Azapa Valley (18°35′ S, 69°30′ W) in northern Chile, with approximately 600 ha of fresh tomatoes under greenhouses. Cultivars resistant to Fusarium oxysporum f. sp. lycopersici (FOL) races 1 and 2 are mainly used. However, in 2012 and 2013, Fusarium wilt incidence was 2 to 3%. Symptoms appeared unilaterally and consisted of yellowing, leaf wilting of lower leaves, dark brown vascular discoloration, and plant death. The aim of this study was to determine the causal agent of tomato wilt in seven tomato greenhouses in the Azapa Valley. Stem samples (5 × 5 mm) were obtained 10 cm of the stem base from wilted tomatoes ‘Naomi’ (BIOAMERICA S.A., Chile) or from Maxifort tomato rootstock (De Ruiter Seed, USA), both FOL resistant to races 1 and 2. Samples were washed with tap water, surface sterilized with 1% NaClO for 3 min, and incubated on sterile moist paper towels in petri plates for 5 days at 22°C. Mycelial fragments from white colonies, emerging from diseased tissues, were transferred to PDA. Six Fusarium isolates were characterized by the presence of hyaline macroconidia, mostly 3 to 5 septate, slightly curved (19.2 to 32.1 × 2.9 to 4.5 μm) and single-celled, oval to elongated microconidia (3.1 to 8.9 × 2.0 to 4.0 μm). Chlamydospores were single or in pairs. These isolates were identified as F. oxysporum (3). The identity of F. oxysporum was confirmed by PCR assays using genomic DNA of each isolated and the universal primers Uni F and Uni R that generate a 672-bp PCR product. The pathogenic form and races were determined by PCR assays using the specific primers uni, sp13, sp23, and sprl that were able to discriminate all the three FOL races as well as F. oxysporum f. sp. radicis-lycopersici (FORL) isolates (2). The sp13 and sp23 primers amplified DNA bands of 445 and 518 bp, confirming the identity of FOL race 3. However, sprl amplified a fragment of 947 bp corresponding to FORL (2). Pathogenicity tests were conducted on 25-day-old seedlings (10 seedlings per isolate) of tomato ‘Poncho Negro,’ which is susceptible to FOL and FORL. Seedling roots were cut, submerged for 5 min in conidial suspension of 2 × 106 conidia/ml, and transplanted to 250-ml plastic containers with sterile substrate (sand/peat, 1:1). Equally treated non-inoculated seedlings were left as controls. The first symptoms induced by each of the five FOL isolates appeared 8 days after incubation under greenhouse and were characterized by yellowing of older leaves, sometimes affecting one side of the plant, vascular discoloration of the stem, and eventually plant death. In contrast, all seedlings inoculated with a FORL isolate developed a necrotic lesion and vascular discoloration at the base of the stems near the soil line, followed by wilting and plant death. Control plants remained asymptomatic. F. oxysporum was re-isolated only from inoculated plants, completing Koch's postulates. FOL and FORL were reported earlier in other tomato growing areas of Chile (1), located over 1,000 km south of the Azapa Valley. However, this is the first report of FOL race 3 and FORL in the Azapa Valley and FOL race 3 is reported for the first time in Chile. References: (1) S. Acuña. Compendio de Fitopatógenos de Cultivos Agrícolas. Servicio Agrícola y Ganadero. Gobierno de Chile, 2008. (2) Y. Hirano and T. Arie. J. Gen. Plant Pathol. 72:273, 2006. (3) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006.
G. Sepúlveda-Chavera, R. Salvatierra-Martínez, and R. Andía-Guardia. 2013. The alternative control of powdery mildew complex (Leveillula taurica and Erysiphe sp.) in tomato in the Azapa Valley, Chile. Cien. Inv. Agr. 40(1):119-130. The powdery mildew complex (Leveillula taurica and Erysiphe sp.) is one of the major factors limiting tomato (Solanum lycopersicum) production in the Azapa Valley, Chile. This problem is especially severe in the case of protected crops because the options for the treatment of these crops with chemical fungicides are limited. This study evaluated the effects of 0.5 and 1.0 kg·100 L -1 Ospo-V55® and 10% raw cow's milk on tomato leaflets infected by powdery mildew complex. Ospo-V55®, with active ingredients consisting of 10% p/p of algae extract (IB-101) and 0.8% of microorganism concentrate, was evaluated in this study. Ospo-V55® contains flavonoids, alkaloids and phenols, oligo-and polysaccharides, 8.6% protein, 0.5% magnesium and trace amounts of riboflavin, niacin and ascorbic acid. Tacora Mas® (a mixture of 25% tebuconazole and 12.5% carbendazime) and Acoidal WG (wettable sulfur) were included for comparison with the Ospo-V55® and raw cow's milk treatments. A diagrammatic scale to assess the severity of the disease was validated and used to quantify the damage resulting from tomato powdery mildew and the effects of the treatments. The effect of the treatments on the germination of the conidia of L. taurica and Erysiphe sp. was also studied on water agar. Tacora Mas® provided the best control of powdery mildew in the field, followed by 1.0 kg·100 L -1 Ospo-V55® and Acoidal. However, Ospo-V55® was more effective than Acoidal. All treatments, including raw cow´s milk, significantly (P≤0.05) reduced disease severity in the leaf compared with the control. Both surfaces of the leaves were assessed, and better results were obtained on the upper surface. This difference was associated with differences in wetting. Acoidal was the most effective treatment for inhibiting conidia germination, decreasing germination more than 98.2%. This result differed significantly from the value of inhibition produced by Tacora Mas®. Ospo-V55® and raw cow´s milk showed significant differences (P≤0.05) in conidial germination compared with the control. Based on these results, we conclude that Ospo-V55 and raw cow´s milk can be used in an integrated management program as alternative treatments to control tomato powdery mildew complex.
During the last decades, the incorporation of beneficial microorganisms in agriculture crop management has become a common practice. Seed coating of these microorganisms still faces technical issues, which limit its implementation in conventional agriculture. An adaption to widely established agricultural practices, e.g. fertigation, could help to overcome these issues. Here, using Bacillus velezensis strain BBC047, we show the influence of the crop phenological stages on the efficiency and success of microbial inoculation under agricultural conditions. In the commercial nursery, strain BBC047 improved growth in a variety of horticulture crops like basil, cabbage, tomato and bell pepper, the latter with the strongest effects in strengthening and accelerating the seedling growth (root and aerial biomass). For a field trial under productive conditions, different application strategies were compared, using bell pepper (Capsicum annuum L.) as crop under fertigation: conventional management (T1), application to the seedling (only nursery, T2), only post-transplant application (field, T3) and a combination of both (T4). In T2 and T4, the post-transplantation survival rate (p < 0.05) improved and the productivity of the plants increased (> 100%). Applications of BBC047 post-transplantation (T3) caused a lower increase in productivity (25%). Fruits from all three application strategies contained significantly more Vitamin C. We conclude that in conventional agriculture, the applications of PGPR inoculants to early crop phenological stages like nurseries are a viable alternative for the efficient use of PGPR inoculants. In comparison, a late introduction of a PGPR reduces its beneficial effect on crop productivity. We highlight that an appropriate timing in the use of PGPR inoculants is crucial for product development and success in sustainable agriculture.
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