M. S. Reddy

Bacterial volatiles promote growth in Arabidopsis

Several chemical changes in soil are associated with plant growth-promoting rhizobacteria (PGPR). Some bacterial strains directly regulate plant physiology by mimicking synthesis of plant hormones, whereas others increase mineral and nitrogen availability in the soil as a way to augment growth. Identification of bacterial chemical messengers that trigger growth promotion has been limited in part by the understanding of how plants respond to external stimuli. With an increasing appreciation of how volatile organic compounds signal plants and serve in plant defense, investigations into the role of volatile components in plant–bacterial systems now can follow. Here, we present chemical and plant-growth data showing that some PGPR release a blend of volatile components that promote growth of Arabidopsis thaliana. In particular, the volatile components 2,3-butanediol and acetoin were released exclusively from two bacterial strains that trigger the greatest level of growth promotion. Furthermore, pharmacological applications of 2,3-butanediol enhanced plant growth whereas bacterial mutants blocked in 2,3-butanediol and acetoin synthesis were devoid in this growth-promotion capacity. The demonstration that PGPR strains release different volatile blends and that plant growth is stimulated by differences in these volatile blends establishes an additional function for volatile organic compounds as signaling molecules mediating plant–microbe interactions.

Bacterial Volatiles Induce Systemic Resistance in Arabidopsis

Plant growth-promoting rhizobacteria, in association with plant roots, can trigger induced systemic resistance (ISR). Considering that low-molecular weight volatile hormone analogues such as methyl jasmonate and methyl salicylate can trigger defense responses in plants, we examined whether volatile organic compounds (VOCs) associated with rhizobacteria can initiate ISR. In Arabidopsis seedlings exposed to bacterial volatile blends from Bacillus subtilis GB03 and Bacillus amyloliquefaciens IN937a, disease severity by the bacterial pathogen Erwinia carotovora subsp. carotovora was significantly reduced compared with seedlings not exposed to bacterial volatiles before pathogen inoculation. Exposure to VOCs from rhizobacteria for as little as 4 d was sufficient to activate ISR in Arabidopsis seedlings. Chemical analysis of the bacterial volatile emissions revealed the release of a series of low-molecular weight hydrocarbons including the growth promoting VOC (2R,3R)-(-)-butanediol. Exogenous application of racemic mixture of (RR) and (SS) isomers of 2,3-butanediol was found to trigger ISR and transgenic lines of B. subtilis that emitted reduced levels of 2,3-butanediol and acetoin conferred reduced Arabidopsis protection to pathogen infection compared with seedlings exposed to VOCs from wild-type bacterial lines. Using transgenic and mutant lines of Arabidopsis, we provide evidence that the signaling pathway activated by volatiles from GB03 is dependent on ethylene, albeit independent of the salicylic acid or jasmonic acid signaling pathways. This study provides new insight into the role of bacteria VOCs as initiators of defense responses in plants.

Induced systemic protection against tomato late blight elicited by plant growth-promoting rhizobacteria.

ABSTRACT Two strains of plant growth-promoting rhizobacteria (PGPR), Bacillus pumilus SE34 and Pseudomonas fluorescens 89B61, elicited systemic protection against late blight on tomato and reduced disease severity by a level equivalent to systemic acquired resistance induced by Phytophthora infestans or induced local resistance by chemical inducer beta-amino butyric acid (BABA) in greenhouse assays. Germination of sporangia and zoospores of P. infestans on leaf surfaces of tomato plants treated with the two PGPR strains, pathogen, and chemical BABA was significantly reduced compared with the noninduced control. Induced protection elicited by PGPR, pathogen, and BABA were examined to determine the signal transduction pathways in three tomato lines: salicylic acid (SA)-hydroxylase transgenic tomato (nahG), ethylene insensitive mutants (Nr/Nr), and jasmonic acid insensitive mutants (def1). Results suggest that induced protection elicited by both bacilli and pseudomonad PGPR strains was SA-independent but ethylene- and jasmonic acid-dependent, whereas systemic acquired resistance elicited by the pathogen and induced local resistance by BABA were SA-dependent. The lack of colonization of tomato leaves by strain 89B61 suggests that the observed induced systemic resistance (ISR) was due to systemic protection by strain 89B61 and not attributable to a direct interaction between pathogen and biological control agent. Although strain SE34 was detected on tomato leaves, ISR mainly accounted for the systemic protection with this strain.

The role of salicylic acid in induced systemic resistance elicited by plant growth-promoting rhizobacteria against blue mold of tobacco

Investigations were conducted to determine the role of salicylic acid (SA) in induced systemic (ISR) resistance against blue mold disease of tobacco elicited by plant growth-promoting rhizobacteria (PGPR). SA did not inhibit germination of sporangia or development of germ tubes of Peronospora tabacina, the blue mold pathogen, in vitro. Of three PGPRstrains tested, none produced detectable levels of SA in aqueous Murashige and Skoog (MS) medium, and only Serratia marcescens strain 90-166 produced SA in tryptic soy broth (TSB). In a microtiter plate system, levels of endogenous free SA in tobacco (Nicotiana tabacum) seedlings treated with three PGPRstrains significantly increased during the first week after PGPRtreatment. In the second week, however, levels of SA were significantly lower than those in nontreated tobacco seedlings. When plants treated with Bacillus pumilus strain SE34 were challenged with P. tabacina, levels of SA increased markedly 1 day after challenge, compared to the nonbacterized and challenged control. However, a similar increase in SA 1 day after pathogen challenge did not occur in plants treated with PGPRstrains 90-166 or Pseudomonas fluorescens strain 89B-61. These observations indicate that SA accumulation in tobacco plants may play a role in ISRagainst tobacco blue mold by PGPR . Disease assays conducted in the microtiter plates showed that the tested PGPRstrains significantly reduced disease severity of blue mold in both Xanthi-nc and transgenic NahG tobacco, indicating that systemically induced resistance in tobacco to blue mold by PGPRmay be SA-independent. 2002 Elsevier Science (USA). All rights reserved.

Rhizobacteria-Mediated Growth Promotion of Tomato Leads to Protection Against Cucumber mosaic virus

ABSTRACT We evaluated combinations of two strains of plant growth-promoting rhizobacteria (PGPR) formulated with the carrier chitosan for the ability to induce growth promotion of tomato plants and resistance to infection by Cucumber mosaic virus (CMV). Each PGPR combination included GB03 (Bacillus subtilis) and one of the following PGPR strains: SE34 (B. pumilus), IN937a (B. amyloliquefaciens), IN937b (B. subtilis), INR7 (B. pumilus), or T4 (B. pumilus). The PGPR combinations formulated with chitosan are referred to as biopreparations. Tomato plants treated with each of the biopreparations appeared phenotypically and developmentally similar to nonbacterized control plants that were 10 days older (referred to as the older control). When plants were challenged with CMV, all plants in the biopreparation treatments and the older control treatment had significantly greater height, fresh weight, and flower and fruit numbers than that of plants in the CMV-inoculated same age control treatment. CMV disease severity ratings were significantly lower for biopreparation-treated and older control tomato plants than for that of same age control plants at 14 and 28 days postinoculation (dpi). CMV accumulation in young noninoculated leaves was significantly less for all biopreparation-treated plants and those in the older control than for the same age control plants at 14 dpi and for four of the five biopreparation treatments at 28 dpi. In those tomato plants shown to be infected, the amount of CMV in noninoculated leaves was significantly lower for three of the biopreparation treatments and the older control treatment at 14 dpi and biopreparation G/INR7 treatment at 28 dpi when compared with the control treatment. These data show that treatment of tomato plants with biopreparations results in significant enhancement of growth and protection against infection by CMV.

Combined application of the biological product LS213 with Bacillus, Pseudomonas or Chryseobacterium for growth promotion and biological control of soil-borne diseases in pepper and tomato

Recent works suggest that the combination of several PGPRs could be more effective than individual strains as a horticultural product. LS213 is a product formed by a combination of two PGPRs, Bacillus subtilis strain GB03 (a growth-promoting agent), B. amyloliquefaciens strain IN937a (an inducer of systemic resistance) and chitosan. The aim of this work is to establish if the combination of three PGPR, B. licheniformis CECT 5106, Pseudomonas fluorescens CECT 5398 and Chryseobacterium balustinum CECT 5399 with LS213 would have a synergistic effect on growth promotion and biocontrol on tomato and pepper against Fusarium wilt and Rhizoctonia damping off. When individual rhizobacterium and the LS213 were put together, the biometric parameters were higher than with individual rhizobacterium both in tomato and pepper, revealing a synergistic effect on growth promotion, being the most effective combination that of B. licheniformis and LS213. When P. fluorescens CECT 5398 was applied alone, it gave good results, which could be due to the production of siderophores by this strain. Biocontrol results also indicate that those treatments that combined LS213 and each of the bacteria (Treatments: T7 and T8) gave significantly higher percentages of healthy plants for both tomato (T7: 65%) and pepper (T7: 75% and T8: 70%) than the LS213 alone (45% of healthy plants for tomato and 60% for pepper) three weeks after pathogen attack. The effects in pepper were more marked than in tomato. The best treatment in biocontrol was the combination of P. fluorescens and LS213. In summary, the combination of microorganisms gives better results probably due to the different mechanisms used.

Comparative performance of formulations of plant growth promoting rhizobacteria in growth promotion and suppression of downy mildew in pearl millet

Five plant growth promoting rhizobacterial formulations, each consisting of two Bacilli strains with chitosan as a carrier were tested for their capacity to promote growth and induce resistance against downy mildew in pearl millet under both greenhouse and field conditions. Three modes of applications were tested: seed treatment, soil amendment, and seed treatment+soil amendment. In general, irrespective of application method, most of the formulations, in comparison with the control, increased plant growth and vigor as measured by seed germination, seedling vigor, plant height, fresh and dry weight, leaf area, tillering capacity, number of earheads, length and girth of earhead, 1000 seed weight and yield. The time of flowering was also advanced by 4–5 days over the control. Likewise all the formulations significantly reduced downy mildew incidence relative to the nontreated control. However, the rate of growth enhancement and disease suppression varied considerably with the formulations. Formulations LS256 and LS257 besides being the best growth promoters were also the most efficient resistance inducers. None of the formulations matched the level of the fungicide metalaxyl in offering protection against downy mildew. Among the application methods tested, soil amendment was found to be the most suitable and desirable way of delivering the formulations. Combination of seed treatment and soil amendment produced the same effect that was produced by soil amendment alone. The study demonstrates a potential role for plant growth promoting rhizobacterial formulations in downy mildew management.

Advances in genetically engineered (transgenic) plants in pest management—an over view

Abstract Transgenic plants are produced via Agrobacterium mediated transformation and other direct DNA transfer methods. A number of transgenes conferring resistance to insects, diseases and herbicide tolerance have been transferred into crop plants from a wide range of plant and bacterial systems. In the majority of the cases, the genes showing expression in transgenic plants are stably inherited into the progeny without detrimental effects on the recipient plant. More interestingly, transgenic plants under field conditions have also maintained increased levels of insect resistance. Now, transgenic crops occupy 44.2 million hectares on global basis. During the last 15 years, transformations have been produced in more than 100 plant species; notable examples include maize, wheat, soybean, tomato, potato, cotton, rice, etc. Amongst these herbicide tolerant and insect tolerant cotton, maize and soybean carrying Bacillus thuringiensis ( Bt ) genes are grown on a commercial scale. Genetic transformation and gene transfer are routine in many laboratories. However, isolation of useful genes and their expression to the desired level to control insect pests still involves considerable experimentation and resources. Developing pest resistant varieties by insertion of a few or single specific gene(s) is becoming an important component of breeding. Use of endotoxin genes such as Bt and plant derived genes (proteinase inhibitors) to the desired levels offers new opportunities to control insects and strategies involving combination of genes. Transgenic technology should be integrated in a total system approach for ecologically friendly and sustainable pest management. Issues related to Intellectual property rights, regulatory concerns, and public perceptions for release of transgenics need to be considered. Providing wealth of information on gene expression in higher plants by switching the gene on and off as and when required, makes gene manipulation a more direct process for genetic improvement of crops.

Lack of Induced Systemic Resistance in Peanut to Late Leaf Spot Disease by Plant Growth-Promoting Rhizobacteria and Chemical Elicitors

A disease assay was optimized for late leaf spot disease of peanut using Cercosporidium per-sonatum in the greenhouse, and this assay was used in attempts to elicit induced systemic resistance using strains of plant growth-promoting rhizobacteria (PGPR) and chemical elicitors. Nineteen strains of spore-forming bacilli PGPR, including strains of Paenibacillus macerans, Brevibacillus brevis, Bacillus laterosporus, B. subtilis, B. pumilus, B. amyloliquefaciens, B. sphaericus, B. cereus, and B. pasteurii, which previously elicited systemic disease control activity on other crops, were evaluated in greenhouse assays. Seven PGPR strains elicited significant disease reduction in a single experiment; however, none repeated significant protection achieved in the greenhouse assay, while significant protection consistently occurred with the fungicide chlorothalonil (Bravo). In other greenhouse trials, neither stem injections of C. personatum nor foliar sprays of chemicals, including salicylic acid, sodium salicylate, isonicotinic acid, or benzo[1,2,3]thiadiazole-7-carbothioc acid S-methyl ester (Actigard), which elicit systemic acquired resistance on other crops, elicited significant disease protection. In contrast, foliar sprays with DL-β-amino-n-butyric acid (BABA), which is an elicitor of localized acquired resistance, resulted in significantly less late leaf spot disease in one of two tests. Combination treatments of four PGPR strains with BABA in the greenhouse did not significantly protect peanut from late leaf spot. Field trials conducted over two growing seasons indicated that none of the 19 PGPR strains, applied as seed treatments at two concentrations, significantly reduced late leaf spot disease. The same chemical elicitors tested in the greenhouse, including BABA, did not elicit significant disease protection. Some combinations of four PGPR and BABA significantly reduced the disease at one but not at two sample times. Collectively, these results suggest that late leaf spot resistance in peanut is not systemically inducible in the same manner as is resistance to diseases in other crops by PGPR and chemical inducers.

Induction of Growth Promotion and Resistance Against Downy Mildew on Pearl Millet (Pennisetum glaucum) by Rhizobacteria

A series of laboratory, greenhouse, and field experiments were conducted to evaluate seven strains of plant growth-promoting rhizobacteria (PGPR). The PGPR were tested as suspensions of fresh cultures and talc-based powder formulations. Evaluations were conducted on pearl millet (Pennisetum glaucum) for growth promotion and management of downy mildew caused by Sclerospora graminicola. All treatments with fresh suspensions and powdered formulations showed enhancement in germination and vigor index over the respective untreated controls. With fresh suspensions, maximum vigor index resulted from treatments by Bacillus pumilus strain INR7 followed by B. subtilis strain IN937b (64 and 38% higher than the untreated control, respectively). With powdered formulation, treatment with strain INR7 also resulted in the highest germination and vigor indexes, which were 10 and 63%, respectively, over the untreated control. Under experimental plot conditions, prominent enhancement in growth also was observed in the disease tests. Yield was enhanced 40 and 37% over the untreated control by seed treatment with powdered formulations of strains INR7 and SE34, respectively. The same strains also increased yield by 36 and 33%, respectively, when applied as fresh suspensions. Studies on downy mildew management resulted in varied degrees of protection by the PGPR both under greenhouse and field conditions. With fresh suspensions, treatment with INR7 resulted in the highest protection (57%), followed by B. pumilus strain SE34 and B. subtilis strain GBO3, which resulted in 50 and 43% protection, respectively, compared with the untreated control. With powdered formulation, PGPR strain INR7 suppressed downy mildew effectively, resulting in 67% protection, while SE34 resulted in 58% protection, followed by GBO3 with 56% protection. Treatment with Apron (Metalaxyl) resulted in the highest protection against downy mildew under both greenhouse and field conditions. Thus, the present study suggests that the tested PGPR, both as powdered formulations and fresh suspensions, can be used within pearl millet downy mildew management strategies and for plant growth promotion.