Induced Resistance by β‐Aminobutyric Acid in Artichoke against White Mould Caused by Sclerotinia sclerotiorum

β‐aminobutyric acid (BABA) was assessed for the ability to protect two artichoke cultivars, C3 and Exploter, against white mould caused by Sclerotinia sclerotiorum, which represents a major problem in the cultivation of this crop in many growing areas of Central Italy. Changes in the activity and isoenzymatic profiles of the pathogenesis‐related (PR) proteins β‐1,3‐glucanase, chitinase and peroxidase in plantlets upon BABA treatment and following inoculation of the pathogen in plantlets and leaves detached from adult plants were also investigated as molecular markers of induced resistance and priming. BABA treatments by soil drenching induced a high level of resistance against S. sclerotiorum in artichoke plantlets of both cultivars C3 and Exploter with a similar level of protection and determined a consistent increase in peroxidase activity paralleled with the differential induction of alkaline isoenzyme with a pI 8.6. A consistent change was found in Exploter in the peroxidase activity following BABA treatments and pathogen inoculation and was paralleled with the expression of an anionic band in plantlets and both anionic and cationic bands in leaves. Our results showed a correlation between BABA‐induced resistance (BABA‐IR) and a augmented capacity to express basal defence responses, more pronounced in cultivar C3 and associated β‐1,3‐glucanase accumulation in both plantlets and leaves inoculated with the pathogen, whereas chitinase resulted affected only at plantlet stage. The present results represent the first one showing the effect of BABA in inducing resistance in artichoke and associated accumulation of selected PRs. If confirmed in field tests, the use of BABA at early plant stages may represent a promising approach to the control soilborne pathogens, such as the early infection of S. sclerotiorum.     

Treating seeds with activators of plant defence generates long-lasting priming of resistance to pests and pathogens

? Priming of defence is a strategy employed by plants exposed to stress to enhance resistance against future stress episodes with minimal associated costs on growth. Here, we test the hypothesis that application of priming agents to seeds can result in plants with primed defences. ? We measured resistance to arthropod herbivores and disease in tomato (Solanum lycopersicum) plants grown from seed treated with jasmonic acid (JA) and/or β-aminobutryric acid (BABA). ? Plants grown from JA-treated seed showed increased resistance against herbivory by spider mites, caterpillars and aphids, and against the necrotrophic fungal pathogen, Botrytis cinerea. BABA seed treatment provided primed defence against powdery mildew disease caused by the biotrophic fungal pathogen, Oidium neolycopersici. Priming responses were long-lasting, with significant increases in resistance sustained in plants grown from treated seed for at least 8 wk, and were associated with enhanced defence gene expression during pathogen attack. There was no significant antagonism between different forms of defence in plants grown from seeds treated with a combination of JA and BABA. ? Long-term defence priming by seed treatments was not accompanied by reductions in growth, and may therefore be suitable for commercial exploitation.     

Accumulation patterns of endogenous β‐aminobutyric acid during plant development and defence in Arabidopsis thaliana

• We recently discovered that β‐aminobutyric acid (BABA), a molecule known for its ability to prime defences in plants, is a natural plant metabolite. However, the role played by endogenous BABA in plants is currently unknown. In this study we investigated the systemic accumulation of BABA during pathogen infection, levels of BABA during plant growth and development and analysed mutants possibly involved in BABA transport or regulation.
• BABA was quantified by LC‐MS using an improved method adapted from a previously published protocol. Systemic accumulation of BABA was determined by analysing non‐infected leaves and roots after localised infections with Plectosphaerella cucumerina or Pseudomonas syringae pv. tomato (Pst) DC3000 avrRpt2. The levels of BABA were also quantified in different plant tissues and organs during normal plant growth, and in leaves during senescence. Mutants affecting amino acid transport (aap6, aap3, prot1 and gat1), γ‐aminobutyric acid levels (pop2) and senescence/defence (cpr5‐2) were analysed.
• BABA was found to accumulate only locally after bacterial or fungal infection, with no detectable increase in non‐infected systemic plant parts. In leaves, BABA content increased during natural and induced senescence. Reproductive organs had the highest levels of BABA, and the mutant cpr5‐2 produced constitutively high levels of BABA.
• Synthetic BABA is highly mobile in the receiving plant, whereas endogenous BABA appears to be produced and accumulated locally in a tissue‐specific way. We discuss a possible role for BABA in age‐related resistance and propose a comprehensive model for endogenous and synthetic BABA.

     

Proteomic study of β-aminobutyric acid-mediated cadmium stress alleviation in soybean

The present study highlights the protective role of β-aminobutyric acid (BABA) in alleviating cadmium (Cd) stress in soybean. Proteomic analyses revealed that out of 66 differentially abundant protein spots in response to Cd challenge, 17 were common in the leaves of BABA-primed and non-primed plants. Oxygen-evolving enhancer protein 1 and ribulose bisphosphate carboxylase small chain 1 were detected in increase abundance in both groups of leaves. Among the 15 commonly decreased protein spots, the relative intensity levels of heat shock cognate 70-kDa protein, carbonic anhydrase, methionine synthase, and glycine dehydrogenase were partially restored after BABA treatment. Moreover, BABA priming significantly enhanced the abundance of the defense-related protein peroxiredoxin and glycolytic enzymes in response to Cd exposure. Additionally, the impact of Cd on the physiological state of BABA-primed and non-primed plants was analyzed using a biophoton technique. The finding of comparatively low biophoton emission in BABA-primed leaves under Cd stress indicates that these plants experienced less oxidative damage than that of non-primed plants. Proteomic study coupled with biophoton analysis reveals that BABA pretreatment helps the plants to combat Cd stress by modulating plants' defence mechanism as well as activating cellular detoxification system to protect the cells from Cd induced oxidative stress damages.     

Soil Drench Treatment with ?-Aminobutyric Acid Increases Drought Tolerance of Potato

The non-protein amino acid β-aminobutyric acid (BABA) is known to be a priming agent for a more efficient activation of cellular defence responses and a potent inducer of resistance against biotic and abiotic stresses in plants. Nevertheless, most of the studies on priming have been carried out in Arabidopsis. In potato, the effect of BABA was demonstrated only on biotic stress tolerance. We investigated the effect of BABA on the drought tolerance of potato and found that soil drenched with BABA at a final concentration of 0.3 mM improves the drought tolerance of potato. Water loss from the leaves of the primed plants is attenuated and the yield is increased compared to the unprimed drought-stressed plants. The metabolite composition of the tubers of the BABA-treated plants is less affected by drought than the tuber composition of the non-treated plants. Nitric oxide and ROS (reactive oxygen species) production is increased in the BABA-treated roots but not in the leaves. In the leaves of the BABA-treated plants, the expression of the drought-inducible gene StDS2 is delayed, but the expression of ETR1, encoding an ethylene receptor, is maintained for a longer period under the drought conditions than in the leaves of the non-treated, drought-stressed control plants. This result suggests that the ethylene-inducible gene expression remains suppressed in primed plants leading to a longer leaf life and increased tuber yield compared to the non-treated, drought-stressed plants. The priming effect of BABA in potato, however, is transient and reverts to an unprimed state within a few weeks.     

Induction of Local and Systemic Resistance to Colletotrichum coccodes in Pepper Plants by dl -β-Amino-n-Butyric Acid

dl -β-amino- n -butyric acid (BABA) that was applied as a foliar spray or soil drench to pepper plants induced local and systemic resistance to a challenge infection with Colletotrichum coccodes . About 85 and 100% protections against the anthracnose were achieved by a relatively high concentration of BABA at 1000 μg/ml, which had no antifungal activity in vitro against C. coccodes . Protection was expressed as a reduction in the number and size of lesions. dl -α-amino- n -butyric acid (AABA) was not so effective as BABA, whereas dl -γ-amino- n -butyric acid (GABA) provided a little protection against anthracnose. At least 5 days were needed after BABA treatment as a soil drench in order to induce resistance in pepper plants. After BABA treatment as a soil drench, the plants remained protected over 15 days. Application of BABA to the lower leaves significantly protected the leaves above the treated leaves from C. coccodes infection, which suggested that systemic resistance to the anthracnose was induced in pepper plants by BABA. The leaves above the BABA-treated lower leaves were strongly protected by reduction of the number and size of lesions.     

Wounding of Arabidopsis leaves causes a powerful but transient protection against Botrytis infection

Physical injury inflicted on living tissue makes it vulnerable to invasion by pathogens. Wounding of Arabidopsis thaliana leaves, however, does not conform to this concept and leads to immunity to Botrytis cinerea , the causal agent of grey mould. In wounded leaves, hyphal growth was strongly inhibited compared to unwounded controls. Wound-induced resistance was not associated with salicylic acid-, jasmonic acid- or ethylene-dependent defence responses. The phytoalexin camalexin was found to be involved in this defence response as camalexin-deficient mutants were not protected after wounding and the B. cinerea strains used here were sensitive to this compound. Wounding alone did not lead to camalexin production but primed its accumulation after inoculation with B. cinerea , further supporting the role of camalexin in wound-induced resistance. In parallel with increased camalexin production, genes involved in the biosynthesis of camalexin were induced faster in wounded and infected plants in comparison with unwounded and infected plants. Glutathione was also found to be required for resistance, as mutants deficient in γ-glutamylcysteine synthetase showed susceptibility to B. cinerea after wounding, indicating that wild-type basal levels of glutathione are required for the wound-induced resistance. Furthermore, expression of the gene encoding glutathione- S -transferase 1 was primed by wounding in leaves inoculated with B. cinerea . In addition, the priming of MAP kinase activity was observed after inoculation of wounded leaves with B . cinerea compared to unwounded inoculated controls. Our results demonstrate how abiotic stress can induce immunity to virulent strains of B. cinerea , a process that involves camalexin and glutathione.     

Priming for enhanced defence responses by specific inhibition of the Arabidopsis response to coronatine

The priming agent β-aminobutyric acid (BABA) is known to enhance Arabidopsis resistance to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) DC3000 by potentiating salicylic acid (SA) defence signalling, notably PR1 expression. The molecular mechanisms underlying this phenomenon remain unknown. A genome-wide microarray analysis of BABA priming during Pst DC3000 infection revealed direct and primed up-regulation of genes that are responsive to SA, the SA analogue benzothiadiazole and pathogens. In addition, BABA was found to inhibit the Arabidopsis response to the bacterial effector coronatine (COR). COR is known to promote bacterial virulence by inducing the jasmonic acid (JA) response to antagonize SA signalling activation. BABA specifically repressed the JA response induced by COR without affecting other plant JA responses. This repression was largely SA-independent, suggesting that it is not caused by negative cross-talk between SA and JA signalling cascades. Treatment with relatively high concentrations of purified COR counteracted BABA inhibition. Under these conditions, BABA failed to protect Arabidopsis against Pst DC3000. BABA did not induce priming and resistance in plants inoculated with a COR-deficient strain of Pst DC3000 or in the COR-insensitive mutant coi1-16. In addition, BABA blocked the COR-dependent re-opening of stomata during Pst DC3000 infection. Our data suggest that BABA primes for enhanced resistance to Pst DC3000 by interfering with the bacterial suppression of Arabidopsis SA-dependent defences. This study also suggests the existence of a signalling node that distinguishes COR from other JA responses.     

Enhanced systemic resistance to bacterial speck disease caused by Pseudomonas syringae pv. tomato by dl-β-aminobutyric acid under salt stress

Plants have evolved different but interconnected strategies to defend themselves against microbial pathogens and stress conditions. The defense responses of tomato (Lycopersicon esculentum Mill) seedlings treated with dl ‐β‐aminobutyric acid (BABA) were investigated with and without abiotic stress (100 mM NaCl) against bacterial speck disease caused by Pseudomonas syringae pv. tomato. The plants were sprayed with 50, 125, 250 or 500 μg ml⁻¹ BABA and were inoculated with 10⁸ colony‐forming units ml⁻¹ bacterial suspension 1 day after treatment. Abiotic stress led to an increase in plant resistance. When BABA was additionally applied as a foliar spray at 125 μg ml⁻¹, the effect on plants was almost identical to that on plants that were sprayed with BABA at 500 μg ml⁻¹ alone. The bacterial multiplication in the plants was 250‐fold lower than in the water‐treated (control) plants and in plants that were sprayed with 500 μg ml⁻¹ BABA alone within 48 h postinoculation (hpi). Physiological studies were carried out in the plants treated with BABA in order to investigate the reason for this synergistic effect. Abiotic stress with BABA spray resulted in high H₂O₂ generation and guaiacol peroxidase activity in the plants. The activity of the enzymatic antioxidative protective system of the plants, superoxide dismutase, ascorbate peroxidase and catalase (CAT), also showed a significant delayed increase in BABA‐treated plants under abiotic stress conditions. These increases in enzyme activity coincided with the initiation of the most suppressive effect of BABA on bacterial growth by 24 hpi, which were significantly higher than the control. Salt stress alone did not lead to any significant increase in CAT activity, but salt stress with BABA did. These findings indicate a synergistic effect between salt stress and BABA at low concentrations, resulting in induced plant resistance. Furthermore, a stress regulation effect of BABA under abiotic stress can be associated with plant resistance.     

Proteomic analysis of β-aminobutyric acid priming and abscisic acid – induction of drought resistance in crabapple (Malus pumila): effect on general metabolism, the phenylpropanoid pathway and cell wall enzymes

In a variety of herbaceous, model and crop plants, DL- β -aminobutyric acid (BABA), has been shown to enhance both biotic and abiotic stress resistance by potentiating rather than inducing resistance responses but studies in woody plants are lacking. In the present study, two-dimensional difference in-gel electrophoresis (DIGE) was used to quantify differences in protein abundance in leaf tissue from BABA-treated, abscisic acid (ABA)-treated and untreated (control) plants during a 10 d drought stress. ABA-treated seedlings were most resistant to water loss followed by BABA-treated seedlings and then untreated seedlings. Although some similarity was observed in the proteome of ABA- and BABA-treated seedlings, a dramatic shift in the proteome occurred earlier in ABA-treated seedlings. Some proteins showed almost identical patterns of increase or decrease in abundance in both BABA- and ABA-treated seedlings, supporting the concept that BABA-induced abiotic stress resistance in plants is achieved by potentiating an ABA-regulated pathway. Some proteins, however, were induced or suppressed only in BABA-primed plants, indicating that BABA may also mediate resistance via some ABA-independent pathways. Based on the putative function of the identified proteins, we propose that changes in cell wall enzymes and a suppression of lignin biosynthesis may play a specific role in BABA-primed drought resistance.     

Resistance phenotypes of transgenic tobacco plants expressing different cucumber mosaic virus (CMV) coat protein genes

Tobacco plants expressing a transgene encoding the coat protein (CP) of a subgroup I strain of cucumber mosaic cucumovirus (CMV), I17F, were not resistant to strains of either subgroup I or II. In contrast, the expression of the CP of a subgroup II strain, R, conferred substantial resistance, but only towards strains of the same subgroup. When protection was observed, the levels of resistance were similar when plants were inoculated with either virions or viral RNA, but resistance was more effective when plants were inoculated with viruliferous aphids. Resistance was not dependent on inoculum strength and was expressed as a recovery phenotype not yet described for plants expressing a CMV CP gene. Recovery could be observed either early in infection (less than one week after inoculation) or later (4 to 5 weeks after inoculation). In plants showing early recovery, mild symptoms were observed on the inoculated leaves, and in some cases symptoms developed on certain lower systemically infected leaves, but the upper leaves were symptomless and virus-free. Late recovery corresponded to the absence of both symptoms and virus in the upper leaves of plants that were previously fully infected. Northern blot analyses of resistant plants suggested that a gene silencing mechanism was not involved in the resistance observed.     

GABA/BABA priming: a means for enhancing abiotic stress tolerance potential of plants with less energy investments on defence cache

Abiotic stress is one of the major factors limiting plant growth and yield globally. Though substantial progress has been made in breeding and genetic manipulation of plants to enhance abiotic stress tolerance, the task remains as a challenge even today. Investigations on the priming activity of various chemicals in plants for enhancing abiotic stress tolerance have been undertaken over the past few years. Priming with γ-amino butyric acid (GABA) and β-amino butyric acid (BABA) gains greater attention, because priming with these non-protein amino acids equips the plants to resist abiotic stresses effectively without suffering costly energy investments in operating defence mechanisms. It is well documented that the protective effect of non-protein amino acids like BABA and GABA on plants is due to a potentiation of natural defence mechanisms against abiotic stresses but at the same time not activating the complete defence arsenal before the stress exposure. The exact mode of action of priming with GABA/BABA in plants is still a puzzle, though their importance as signaling molecules during stress is undoubtful. The better understanding of molecular, physiological, and ecological aspects of GABA/BABA priming might lead to the emergence of this technique as a successful strategy for enhancing the abiotic stress(es) tolerance potential of plants in the field, without compromising much on productivity.     

The effects of a plant defence priming compound, β-aminobutyric acid,on multitrophic interactions with an insect herbivore and a hymenopterous parasitoid

Biocontrol of aphids by natural enemies is utilized in many organic and integrated pest management schemes. β-aminobutyric acid (BABA), a non-protein amino acid, is a plant defence primer that suppresses growth of some insect herbivores when applied as a root drench. This investigation examined how applying BABA to host plants via the roots may impact on a parasitoid wasp of aphids. Female Aphidius ervi (Haliday) did not discriminate against pea aphids (Acyrthosiphon pisum (Harris)) reared on BABA-treated beans (Vicia faba L.) or show any modified responses to volatiles released from BABA-treated plants. BABA reduced the size of emerging wasps, primarily by inhibiting the growth of the host aphid. Metabolomic analysis revealed BABA in both aphids and emergent wasps indicating some potential for direct physiological inhibition to have occurred. Survival of the parasitoids was only reduced at doses of BABA likely to produce phytotoxic effects in many plant species, thus there may be potential to incorporate plant defence primers like BABA into integrated pest management practices. However, the precise mechanisms of BABA-inhibition of insects still require elucidation.     

The effect of β-aminobutyric acid on the growth of herbivorous insects feeding on Brassicaceae

dl ‐β‐Aminobutyric acid (BABA) is a nonprotein amino acid that can enhance defences in a variety of plants against a wide range of pathogens. BABA can also reduce infestation by phytopathogenic nematodes and has recently been shown to suppress the growth of aphids feeding on legumes. This investigation examined the effect of applying BABA as a root drench to a range of Brassicaceae, including Arabidopsis thaliana, on the performance of two species of aphid (Myzus persicae and Brevicoryne brassicae) and the larvae of two species of Lepidoptera (Trichoplusia ni and Plutella xylostella). Application of BABA reduced the performance of all four insect species, and inhibition of insects occurred on all the plants tested. The results illustrate that BABA‐induced resistance (BABA‐IR) can affect generalist and specialist insect herbivores and inhibit insects feeding with mandibulate as well as sap‐feeding mouthparts. The BABA‐induced suppression of B. brassicae and P. xylostella feeding on A. thaliana provides a means to further examine the mechanisms of BABA‐IR to insects using this model plant.     

β-Aminobutyric Acid-induced Resistance in Cucumber against Biotrophic and Necrotrophic Pathogens

The non-protein amino acid β-aminobutyric acid (BABA) protects plants against a wide range of pathogens. Protection of cucumber plants by BABA depends on the potentiation of pathogen specific defence responses. To contribute to the analyses of the mode of action of BABA, we established a protocol for a fast and reproducible leaf disc assay to evaluate the effect of this chemical compound on cucumbers infected with either the biotrophic downy mildew fungus Pseudoperonospora cubensis, or the necrotrophic microbial pathogen Colletotrichum lagenarium . Accumulation of callose could be found in interactions with both pathogens after BABA-treatment. Furthermore, a localized rapid cell death and the production of reactive oxygen intermediates were detected after downy mildew attack. In contrast to this, degenerated primary hyphae were found in BABA induced tissue after inoculation with C. lagenarium .     

Comparative proteomic analysis of BTH and BABA-induced resistance in pea (Pisum sativum) toward infection with pea rust (Uromyces pisi)

Systemic acquired resistance (SAR) to Uromyces pisi in pea was studied by using a proteomic approach. Two-dimensional electrophoresis (2-DE) was used in order to compare the leaf proteome of two pea genotypes displaying different phenotypes (susceptible and partial resistance to the fungus), and in response to parasite infection under the effect of two inducers of SAR, BTH and BABA. Multivariate statistical analysis identified 126 differential protein spots under the experimental conditions (genotypes/treatments). All of these 126 protein spots were subjected to MALDI-TOF/TOF mass spectrometry to deduce their possible functions. A total of 50 proteins were identified using a combination of peptide mass fingerprinting (PMF) and MSMS fragmentation. Most of the identified proteins corresponded to enzymes belonging to photosynthesis, metabolism, biosynthesis, binding and defense response, whose behavior pattern was different in relation to susceptibility/resistance of the genotypes studied and to the BTH/BABA induction to pathogen response. Results obtained in this work suggested that plants could reduce their photosynthesis and other energy metabolism and enhance the production of defense-related proteins to cope the stress. On the other side, we postulated that resistance induced by the chemicals operates via different mechanisms: BABA inducer could act via phenolic biosynthesis pathway, whereas resistance provided by BTH inducer seems to be mediated by defense and stress-related proteins. The results are discussed in terms of response to rust under the effect of inducers.