Induction of total phenolics and defence-related enzymes during β-aminobutyric acid-induced resistance in Brassica carinata against Alternaria blight

Exogenous foliar application of β-aminobutyric acid (BABA) led to a significant reduction in disease severity in Brassica carinata caused by Alternaria brassicae. To get a better insight about changes in defence-related enzymes like phenylalanine ammonia lyase (PAL) and polyphenol oxidase (PPO), isoform analysis of superoxide dismutase (SOD) and peroxidase (POX) were studied. BABA-treated plants showed a significant increase in PAL, PPO enzyme activities and total phenolic content in response to pathogen inoculation. However, isoform analysis of SOD and POX revealed no change in isoform number but a quantitative change in activity was observed in response to pathogen.     

Effect of methyl jasmonate (MeJA) on biochemical responses of wheat seedlings infected by Fusarium culmorum

Fusarium culmorum is a ubiquitous soil-borne fungus which is able to cause foot and root rot on different small-grain cereals, in particular wheat and barley. Wheat (Triticum aestivum L.) is among the oldest and most extensively grown of all crops. Systemic acquired resistance (SAR) is a pathogen-induced disease resistance response in plants that is characterized by broad spectrum disease control. SAR is an important component of the disease resistance repertoire of plants. The effect of the resistance inducer, methyl jasmonate (MeJA), was tested by seed-soaking to determine whether this resistance inducer controlled root rot of wheat caused by F. culmorum. MeJA is an inducing disease resistance in a number of dicotyledonous and monocotyledonous plant species. This research has been conducted to determine the effect of MeJA seed treatment on chlorophyll a, chlorophyll b, total protein, peroxidase (POD) enzyme, phenolic compounds and phenylalanine ammonia lyase (PAL) factors. MeJA, which is an important chemical inducer of SAR, can activate resistance to pathogen infection in commercial crops. Seeds treated with MeJA showed an elevation in the total chlorophyll, carotenoid and activity of defence-related enzymes, POD and PAL.     

Rhizobacterial bioformulation for the effective management of Macrophomina root rot in mungbean

Abstract

Fluorescent pseudomonads based bioformulation was evaluated for their ability to control Macrophomina root rot disease in mungbean (Vigna mungo). P. fluorescens isolate Pf1 showed the maximum inhibition in mycelial growth of Macrophomina phaseolina under in vitro conditions. Bioformulation of Pf1 with chitin was effective in reducing the root rot incidence in green gram both under glasshouse and field conditions. The rhizosphere colonization of P. fluorescens was observed appreciable with the green gram plants. However, Pf1 amended with chitin colonized effectively. Furthermore, the induction of defence-related enzymes and chemicals in plants by Pf1 amended with or without chitin and neem were tested. Increased accumulation of defence enzymes viz., phenylalanine ammonia lyase (PAL), peroxidase (PO), polyphenol oxidase (PPO), chitinase, β-1,3-glucanse and phenolics were observed in Pf1 bioformulation amended with chitin, pre-treated plants challenge inoculated with M. phaseolina under glasshouse conditions. The present study reveals that in addition to direct antagonism and plant-growth promotion, PGPR strains amended with chitin bioformulation induced defence-related enzymes and pathogenesis related (PR) proteins which collectively enhance the resistance in green gram against the infection of M. phaseolina.     

Efficacy of combined applications of antagonist bacteria and chemical resistance inducers for the management of Fusarium solani causing root rot in Withania somnifera

Application of Thiosalicylic acid+Bacillus cereus; O-Acetylsalicylic acid+Pseudomonas fluorescens reduced root rot severity by 85 and 88% and enhanced root yields by 358 and 419%, respectively, against Fusarium solani induced root rot disease in Withania somnifera. Reduction in disease severity was correlated with defence-related enzymes peroxidase, polyphenol oxidase and phenyl ammonium lyase.     

Comparative studies on the activities of chitinase, β-1,3-glucanase, peroxidase and phenylalanine ammonia lyase in the leaves of triticale and wheat infected with Stagonospora nodorum

The activities of chitinase, β-1,3-glucanase, peroxidase and phenylalanine ammonia lyase, constitutive and induced by Stagonospora nodorum were examined in the 10 – 14 day old seedlings of three triticale and two wheat cultivars under controlled environmental conditions and in flag leaves of two triticale cultivars in the field. Two S. nodorum isolates of different virulence were used. Both the constitutive and induced activities in triticale and wheat depended on genotype and in triticale the effect of growth conditions was also evidenced. The constitutive activities of chitinase, β-1,3-glucanase and peroxidase were several fold lower in flag triticale leaves in plants from the field than in the seedlings, growing under controlled conditions, but induction in the infected flag leaves was significantly more pronounced. In triticale genotypic differences in the response to infection were revealed only upon inoculation by S. nodorum isolate of higher virulence. The enzymatic activities increased several fold during successive days after the infection except for phenylalanine ammonia lyase. Induction of this enzyme was only transient and the activity decreased 48 or 96 h after infection when the activities of other enzymes were rising. In flag leaves in the field this activity was differentiated only after infection with more a virulent strain. A tendency appeared in triticale seedlings for association of the resistance to the pathogen with lower enzymatic constitutive activities. This relationship became more evident in triticale infected by S. nodorum and may imply that although the investigated enzymes are certainly involved in general, non-specific defense mechanism, they do not decide on the resistance to pathogen at least in the early stages of infection and cooperate with other factors in the complex pathogen-plant interaction. One can also assume that the enzymatic activities are associated with severity of infection rather than resistance to pathogen.     

Activity profile of defence-related enzymes in rice genotypes (Oryza sativa L.) against rice blast (Magnaporthe oryzae)

The control and infected leaf samples of blast resistant and susceptible rice genotypes were evaluated for activities of defence-related enzymes viz., total phenol content, chitinase, phenylalanine ammonia lyase (PAL), β-glycosidase, antioxidative enzymes, superoxide dismutase, peroxidase and ascorbate peroxidase. The level of total phenol content and the activity profile of chitinase, PAL and β-glycosidase significantly increased in both blast-resistant and susceptible rice genotypes with comparatively higher level induction Tetep, NLR-20104 and Swarnadhan the blast-resistant genotypes. The antioxidative enzymes were comparatively higher in the leaf samples of blast-resistant genotypes recording highest increase in NLR-20104 and KJT-5. The constitutive levels of total phenols and activity of defence-related and antioxidative enzymes in the control leaf samples differed among the genotypes and were even higher in the two blast susceptible genotypes (EK-70 and Chimansal). However, the level of induction as evident from the activity profile differences between control and infected leaf samples suggests higher level of induction was more which is indicative of the induced defence response. The genotype recording maximum induction of defence-related and antioxidative enzymes activity could be useful criteria in screening for blast resistant genotype in rice.     

Role of Phenylalanine Ammonia Lyase and Polyphenol Oxidase in Host Resistance to Bacterial Wilt of Tomato

Plants respond to bacterial pathogen attack by activating various defence responses, which are associated with the accumulation of several factors like defence-related enzymes and inhibitors which serve to prevent pathogen infection. The present study focused on the role of the defence-related enzymes phenylalanine ammonia lyase (PAL) and polyphenol oxidase (PPO) in imparting resistance to tomato against bacterial wilt pathogen Ralstonia solanacearum . The temporal pattern of induction of these enzymes showed maximum activity at 12 h and 15 h for PAL and PPO, respectively, after the pathogen inoculation (hpi) in resistant cultivars. Twenty different tomato cultivars were analyzed for PAL, PPO and total phenol content following pathogen inoculation. The enzyme activities and total phenol content increased significantly (P < 0.05) in resistant cultivars upon pathogen inoculation. The increase in enzyme activities and total phenol content were not significant in susceptible and highly susceptible cultivars. The role of PAL and PPO in imparting resistance to tomato against bacterial wilt disease is discussed.     

Biochemical defence responses of resistant and susceptible rice genotypes against blast pathogen Magnaporthe oryzae

Abstract

Rice blast is the leading fungal disease which is caused by Magnaporthe oryzae that contributes for the significant decline in the rice yield throughout the globe. There is a need for the understanding of biochemical changes in rice plant during blast infection for the development of novel disease control strategies. In the present study, we isolated M. oryzae from the local paddy fields and the fungal isolates (VCF and PON) were identified by ITS-PCR using genomic DNA samples. Further, we inoculated resistant (BR2655 and TUNGA) and susceptible (INTAN and HR12) rice cultivars with PON and VCF isolates. PON isolate showed relatively high virulence compared to VCF and standard MTCC fungal strains. Therefore, we evaluated the effect of PON on the total protein content and plant defence-related key enzymes (peroxidase, polyphenol oxidase, phenylalanine ammonia lyase, β-glucosidase, chitinase and lipoxygenase) activities between 24- and 120-hour post-inoculation (hpi). The results demonstrated the decrease in total protein content in all the inoculated cultivars. In addition, we observed the variation in the activity of peroxidase, polyphenol oxidase, β-glucosidase, chitinase and lipoxygenase at different time points in all the tested rice plants compared to respective controls. However, no significant difference was observed in the phenylalanine ammonia lyase activity relative to its control. Taken together, this study emphasizes on the variation in the activities of plant defence enzymes in different plant cultivars against the tested fungal pathogen and also implementation of defence enzymes as biochemical markers for resistant breeding.     

Defence-related enzymatic response in cabbage to Xanthomonas campestris pv. campestris

Black rot of cabbage caused by Xanthomonas campestris pv. campestris is one of the most important diseases of crucifers worldwide. Expression of defence-related enzymes in cabbage in response to X. campestris pv. campestris was investigated in the current experiment. Among the defence-related enzymes (phynylalanine ammonia lyase, peroxidase, polyphenol oxidase, superoxide dismutase [SOD] and chitinase) and quantity of phenolic compounds studied in the present investigation, phenylalanine ammonia lyase (PAL), the key enzyme in the phenylpropanoid pathway was the first enzyme suppressed at three days after inoculation in X. campestris pv. campestris-cabbage system. Correlation analysis indicated that PAL and phenolic compounds are the two most important compounds determining the susceptibility of cabbage to X. campestris pv. campestris. Induction of peroxidase isoform-1 (Rf value: 0.059) and SOD isoform-1 (Rf value: 0.179) three days after pathogen inoculation implicated the role of these isozymes in susceptible cabbage – X. campestris pv. campestris interaction. This study demonstrates the susceptibility of cabbage to X. campestris pv. campestris is a result of declination of PAL and phenolic contents at biochemical level as a manifestation of increase in bacterial population at the cellular level within the host tissues.     

Induction of defence response against blister blight by calcium chloride in tea

Tea, the second most consumed beverage after water in the world, is produced from the processed tender leaves of tea plants (Camellia sinensis). Production of tea is hindered by various biotic and abiotic stresses. Among the biotic factors, blister blight disease caused by an obligate fungal pathogen, Exobasidium vexans Massee, is a serious problem to the tea industry. The present study was to evaluate the efficacy of elicitor calcium chloride (CaCl₂) in inducing resistance in tea plants against blister blight disease. During the pick time of blister incidence at Darjeeling tea garden, the application of CaCl₂ at a concentration of 1% resulted in disease inhibition around 71% over the control set. Treatment also resulted in the induction of defence enzymes like peroxidase, polyphenol oxidase, phenylalanine ammonia lyase, β-1,3-glucanase and higher phenol accumulation. Furthermore, the increase in defence molecules also correlated with increase in nitric oxide (NO) generation, a potent defence molecule in plant defence. The result suggests that CaCl₂ can used as a potential elicitor in the integrated disease management in organic tea cultivation.     

Induction of systemic acquired resistance in Zea mays L. by Aspergillus flavus and A. parasiticus derived elicitors

Host defence mechanisms can be elicited by using different elicitors produced from the pathogen/host. In this study, an effort has been made to study the effect of two fungal elicitors derived from Aspergillus flavus and A. parasiticus on induction of various defence-related enzymes in maize (Zea mays L.). Foliar application was done on 20-days-old maize plant with 10% A. flavus fungal culture filtrate (AFFCF) and A. parasiticus fungal culture filtrate (APFCF) as elicitors to trigger systemic acquired resistance (SAR). As a response of SAR, an increase in activities of phenylalanine ammonia lyase (PAL), peroxidase (POX), β-1,3-glucanase, nitrate reductase (NR) and nitrite reductase (NiR), total proteins were found highest on 4th day after treatment (DAT), whereas total carbohydrate and total chlorophyll on 2nd and 6th DAT, respectively, in comparison with the control plants. The SDS PAGE analysis revealed the induction of PR proteins, namely Chitinase (25, 29?kDa) and β-1,3-glucanase (33?kDa), in treated plants in comparison with untreated control plants. The treated plants showed enhanced growth and development as well as increase in yield. About 100% survival rate was found in maize seeds treated with AFFCF and APFCF and grown on respective fungal infested soil than control. The enhanced activities of defence enzymes and elevated protein, carbohydrate, chlorophyll content in treated maize plants suggest the induction of SAR against A. flavus and A. parasiticus by using the same fungal elicitors.     

Induction of defence-related enzymes in tomato (Solanum lycopersicum) plants treated with Bacillus subtilis CBR05 against Xanthomonas campestris pv. vesicatoria

Bacterial spot disease caused by Xanthomonas campestris pv. vesicatoria is one of the most important destructive diseases of tomato in many parts of the agricultural world. Therefore, the present study aims to determine the effects of Bacillus subtilis CBR05 inoculation on bacterial spot disease severity and the induction of defence-related enzymes response in tomato. Tomato leaves were evaluated to determine the activities of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and polyphenol oxidase (PPO)) and the content of malondialdehyde (MDA). A reduction in bacterial spot severity was observed in plants inoculated with B. subtilis, compared with those of uninoculated controls. A significant increase in SOD, CAT, POD, and PPO activities was observed in plants treated with B. subtilis after 24?h inoculation compared with non-inoculated pathogen control and mock-inoculated controls. Moreover, the MDA content was induced by pathogen infection, and its amount in B. subtilis inoculated plants was significantly lower than that in pathogen control. Our results suggest that early increases in antioxidant enzymes and the reduction in MDA content with B. subtilis inoculation may play a pivotal role in mitigating oxidative stress, thereby induced systemic resistance against bacterial spot disease in tomato.     

Defence-related Enzymes in Pepper Roots During Interactions with Arbuscular Mycorrhizal Fungi and/or Verticillium dahliae

Previous studies have described that arbuscular mycorrhizal fungi (AMF) can reduce the deleterious effect of Verticillium dahliae Kleb. on pepper growth and yield. In mycorrhizal plants, the bioprotection against soil-borne pathogens can result from the preactivation of defence responses that include some structural modifications and the accumulation of Pathogenesis-Related (PR) proteins. Our first objective was to study if V. dahliae induced defence mechanisms in roots before infected pepper developed visible symptoms of disease. The second aim was to determine if AMF induced defence-related enzymatic activities in pepper roots before or after pathogen’s attack. Results showed that the colonization of pepper roots by Glomus deserticola (Trappe, Bloss and Menge) induced the appearance of new isoforms of acidic chitinases, superoxide dismutase (SOD) and, at early stages, peroxidases. In contrast, V. dahliae neither stimulated the phenylpropanoid pathway nor elicited hydrolytic activities in infected pepper roots. Only in mycorrhizal plants, the inoculation with V. dahliae slightly increased both phenylalanine ammonia-lyase (PAL) and peroxidase activities two weeks later. Mycorrhizal-specific induction of new isoforms of acidic chitinases and SOD together with enhanced peroxidase and PAL activities 2 weeks after pathogen inoculation could be involved in the biocontrol of Verticillium-induced wilt in pepper by AMF.     

Induced defense-related proteins in soybean (Glycine max L. Merrill) plants by Carnobacterium sp. SJ-5 upon challenge inoculation of Fusarium oxysporum

The aim of the present study was to analyze induced expression of defense-related proteins in the soybean plants by rhizobacterial stain Carnobacterium sp. SJ-5 upon challenge inoculation with Fusarium oxysporum. Determination of the enzymatic activity of the different defense-related enzymes, phenylalanine ammonia lyase (PAL), lipoxygenase (LOX), peroxidase (POD) and polyphenol oxidase (PPO) was performed in the major parts of Glycine max L. Merrill using spectrophotometric method. Native-polyacrylamide gel electrophoresis analysis of the POD and PPO was employed followed by activity staining to find out the isoforms of respective enzymes. Activities of the PAL, LOX, POD and PPO were found to be highest in the bacterized root tissue of the soybean plants challenged with F. oxysporum. Isoform analysis revealed that PPO1, PPO4 and POD2 isoforms were expressed at higher levels in bacterized soybean root tissues challenge inoculated with the pathogen. Conclusively it was found that bacterial strain Carnobacterium sp. SJ-5 protect soybean plants from wilt disease caused by F. oxysporum by elicitation of the defense-related enzymes.     

Enhanced Defence Responses of Chickpea Plants Against Rhizoctonia solani by Pre‐Inoculation with Rhizobia

The aim of this work was to study the antagonist effect of two Rhizobium strains Pch Azm and Pch S.Nsir2 to Rhizoctonia solani and for an evaluation of the relative impact of rhizobia on the expression of the plant's defence response against Rhizoctonia. First, these strains reduced fungal growth observed in vitro using the same or separately Petri dishes. Moreover, these isolates led to reduced chickpea infection by R. solani, resulting from the direct effect of rhizobia on pathogens and possible induced resistance in chickpea. Concomitantly, reduction in infection was accompanied by enhanced level of defence‐related enzymes, phenylalanine ammonia lyase (PAL) and peroxidase (POX). An increased level of phenol content was recorded in the roots of bacterized plants grown in the presence of pathogen. The results promise the use of rhizobia for protection of chickpea against R. solani.     

Inducible Proteins in Citrus Rootstocks with Different Tolerance Towards the Root Rot Pathogen Phytophthora palmivora

Activities of defence‐related proteins (β‐1,3‐glucanases, chitinases and peroxidases) and concentrations of total soluble phenolics were measured in roots and leaves of non‐infected and infected plants to investigate the response of different citrus rootstock genotypes to the root rot pathogen Phytophthora palmivora Butler. Infection with the pathogen increased concentrations of total proteins, total phenolics and β‐1,3‐glucanase activity in roots of all genotypes, and increases were associated with the extent of root mass reductions and thus susceptibility of the plants. Root chitinase and root peroxidase levels were slightly reduced or unaltered upon infection. β‐1,3‐Glucanase activity was also elevated in leaves of infected plants, but increases did not differ between tolerant and susceptible rootstocks. Effects of root infection on leaves were typically the reverse of effects on roots for chitinase‐ and peroxidase levels and more pronounced in susceptible rootstock genotypes. Although differences in enzyme expression were observed between susceptible and tolerant citrus seedlings, effects were usually associated with disease progression, and not with resistance to P. palmivora. It is suggested that increased activities of the proteins and soluble phenolics studied are not implicated in the primary defence to Phytophthora root diseases, but may contribute to the inhibition of the pathogen during infection in tolerant citrus.     

Effect of phosphate and the arbuscular mycorrhizal fungus Glomus intraradices on disease severity of root rot of peas (Pisum sativum) caused by Aphanomyces euteiches

 The effects of inorganic phosphate levels and the presence of arbuscular mycorrhiza on disease severity of Aphanomyces euteiches in pea roots were studied. Disease severity on roots and epicotyl as well as the oospore number within infected root tissue were correlated with the phosphorus (P) level in the growth medium. The arbuscular mycorrhizal fungus Glomus intraradices increased P uptake and the P concentration in the plant but reduced disease development in peas. Polyacrylamide gel electrophoresis followed by densitometry of glucose-6-phosphate dehydrogenase specific to A.euteiches was used to measure the activity of the pathogen in roots. The enzyme activity increased with disease severity and disease incidence, except in plants supplemented with P at the highest level, where a peak in activity was seen 12 days after inoculation with the pathogen, followed by a decrease in activity. The epicotyl of mycorrhizal plants showed a reduction in disease severity although this part of the plants was not mycorrhizal. Thus, an induced systemic factor may be responsible for increased resistance in mycorrhizal plants. Accepted: 21 August 1998     

Expression of transferred genes during hairy root development in pea

Root border cell development and expression of reporter genes were evaluated in transgenic pea hairy roots. Successful induction of hairy roots in pea is conditioned by bacterial strain and plant genotype, as well as by developmental and environmental factors. Morphological changes sometimes occur when hairy roots are transferred from infected plants to tissue culture media, but such changes are confined to specific clones. Expression of reporter genes under the control of promoters from bean (Phaseolus vulgaris L.) stress genes encoding phenylalanine ammonia lyase and chalcone synthase were evaluated. Expression patterns vary between hairy roots taken directly from infected plants, and those grown in culture; most hairy roots taken from infected plants exhibit expression throughout all tissues, whereas expression in cultured hairy roots is most often localized to specific tissues. Patterns of expression that occur during different stages of hairy root development are very similar to those observed in transgenic plants expressing the same fusion genes. Border cell separation and release in hairy roots is normal, and expression of glucuronidase in border cells of some transgenic roots resulted in development of bright blue single cells. Cultured hairy roots should provide a very useful model for studying the effect of defined changes in root border cells on microbial associations with roots of this important legume.Abbreviations YEM yeast extract-mannitol - GUS glucuronidase - PAL phenylalanine ammonium lyase - CHS chalcone syntase     

Evaluation of Ochrobactrum anthropi TRS-2 and its talc based formulation for enhancement of growth of tea plants and management of brown root rot disease

Aim: To evaluate Ochrobactrum anthropi TRS‐2 isolated from tea rhizosphere and its talc based formulation for growth promotion and management of brown root rot disease of tea. Methods and Results: Ochrobactrum anthropi TRS‐2, isolated from tea rhizosphere could solubilize phosphate, produce siderophore and IAA in vitro and also exhibited antifungal activity against six test pathogens. Application of an aqueous suspension of O. anthropi to the rhizosphere of nursery grown tea seedlings of five varieties of tea (TV‐18, T‐17, HV‐39, S‐449, UP‐3 and) led to enhanced growth of the treated plants, as evidenced by increase in height, in the number of shoots and number of leaves per shoot. Treatment with O. anthropi also decreased brown root rot of tea, caused by Phellinus noxius. Multifold increase in activities of chitinase, β‐1,3‐glucanase, peroxidase and phenylalanine ammonia lyase in tea plants was observed on application of O. anthropi to soil followed by inoculation with P. noxius. A concomitant increase in accumulation of phenolics was also obtained. Further, talc based formulation of O. anthropi was prepared and its survival determined every month up to a period of 12 months. Ochrobactrum anthropi could survive in the formulation up to a period of 9 months with a concentration of 7·0 log₁₀ CFU g^?1, after which there was a decline. Talc formulation was as effective as aqueous suspensions in both plant growth promotion and disease suppression. Conclusion: Ochrobactrum anthropi, either in aqueous suspension or as talc formulation induced growth of tea plants and suppressed brown root rot disease. It induced defense responses in tea plants. Significance and Impact of the Study: Ochrobactrum anthropi and its talc based formulation can be considered as an addition to available plant growth promoting rhizobacteria (PGPR) currently being used for field application. The present study offers a scope of utilizing this bacterium for growth promotion and disease management which would help in reduction of the use of chemicals in tea plantations.