Growth and physiological and biochemical responses of enset (Ensete ventricosum (Welw.) Cheesman) clones infected with bacterial wilt: Induction of resistance using medicinal plant leaf extract

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Date

2016-06

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Addis Ababa Universty

Abstract

Enset (Ensete ventricosum (Welw.) Cheesman) is a perennial staple food crop widely cultivated in the south and southwestern Ethiopia. It supports the lives of approximately 20 million people. However, its cultivation is impeded by enset bacterial wilt (EBW). Therefore, the aim of the study was to evaluate the antibacterial activity of medicinal plant extracts and antibiotics against Xanthomonas campestris pv. musacearum (Xcm); to study induction of resistance in enset by leaf extract and evaluate its contribution in controlling enset bacterial wilt disease and to investigate growth and metabolic responses of enset clones under different treatments.The antibacterial activities of methanol leaf extracts of Achyranthes aspera, Agarista salicifolia, Datura stramonium, Melia azedarach, Pycnostachys abyssinica and Vernonia amygdalina and some antibiotics; amoxicillin, cephalexin, chloramphenicol, streptomycin sulphate and tetracycline were evaluated in vitro. Qualitative analysis of leaf extracts was carried out using standard methods. Total phenolics content was estimated with Folin Ciocalteau method. The antibacterial activity of methanol leaf extracts was evaluated by disc diffusion method. Minimum inhibitory concentration was determined using agar dilution method. The potential role of A. salicifolia leaf extract to trigger induced resistance, metabolic changes in resistant and susceptible enset clones and the subsequent suppression of EBW was evaluated. Growth, photosynthetic and biochemical responses of the two enset clones, Viz., resistant (Genticha) and susceptible (Midasho) were evaluated 120, 270 and 360 days after treatment application. The phytochemical screening results revealed the existence of alkaloids, phenols, terpenoids, saponnins, tannins and glycosides in extract of A. salicifolia in abundant amount. Similar result was observed in P.abyssinica extract. However, glycosides were absent. Moreover, A. salicifolia leaf extract had the highest total phenolics content. It showed a 100, 85.7, 100, 61.2 and 12.3 % increment from A. aspera, A. salicifolia, D. stramonium, M. azedarach, P. abyssinica and V. amygdalina. The widest inhibition zone diameter was recorded in A. salicifolia extract followed by P. abyssinica. A statistically significant difference in inhibition zone diameter was seen between species and among serial concentrations within a species extract. The two species extracts also showed the lowest minimum inhibitory (MIC) and minimum bactericidal concentrations (MBC). Similarly, antibiotics showed antibacterial activity against Xcm. However, the inhibition zone diameters of antibiotics were significantly higher fivefold times than leaf extracts. The highest anti-Xcm activity was shown by streptomycin sulphate (50.3 mm at 50 mg/mL) followed by amoxicillin (49.7 mm at 50 mg/mL). However, the lowest MIC (0.02 mg/mL) and MBC (0.049 mg/mL) values of antibiotics were recorded by tetracycline. In addition, extract treated and Xcm inoculated plants of both clones had an increased activity of phenylalanine ammonia lyase (PAL), peroxidase, polyphenol oxidase and greater amount of total phenolics content (TPC) and total soluble sugars (TSS). Induced plants of the resistant clone showed an 8, 4.6, 3, 13 and 4.6 fold increments in PAL activity compared to controls at 24, 48, 72, 96 and 120 h post treatment, respectively. Infected control plants on the other hand demonstrated a 7, 3.6, 2.3, 10.6 and 2.4 fold increments compared to controls. Similarly, induced plants of the susceptible clone exhibited 1.5, 1.7, 1.4, 4.2 and 4.4 fold increments in PAL activity compared to controls at 24, 48, 72, 96 and 120 h post treatment, respectively. While infected control plants showed a 0, 1.3, 1.4, 3 and 2 fold increments in PAL activity compared to controls at similar sampling time. The activity of tested enzymes and amounts of TPC and TSS were significantly increased in plants of the resistant clone treated with leaf extract and Xcm as compared to plants of the susceptible clone. The results of the study suggested that the difference between the two clones was partially caused by variations in biochemical responses. Extract induced resistance brought a 33.33 % and 3.1% disease incidence reduction in plants of the susceptible and resistant clones, respectively, compared to infected controls. Plants of the two enset clones also showed variations in growth and physiological responses among themselves and between treatments at different measuring time periods. Nine months after treatment application, induced plants of the resistant clone had significantly lower pseudostem (22.2%), total leaf area (6.8%) and leaf area index (32%) than control plants. Similarly, infected control plants of the susceptible clone showed lower pseudostem girth (29.4%), total leaf area (37.3%) and leaf area index (37.3%) as compared to controls. Generally, the results suggest that extracts of A. salicifolia and P. abyssinica have high potency and the potential for further formulation development as biocides with broad spectrum activity. Tetracycline with the lowest MIC and MBC values is also a potential alternative in controlling EBW. Moreover, the results also suggest the feasibility of induced resistance to control EBW. However, further research should be conducted to formulate the extracts, identify the active compounds responsible for the antibacterial activity of extracts, and evaluate the performance of other application methods under field conditions. Keywords: Agarista salicifolia, Antibiotics, Assimilation rate, Inhibition zone, Ensete ventricosum, Induced resistance, methanol extracts, phenylalanine ammonia lyase, Total phenolic compounds. Xanthomonas campestris pv. musacearum

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Keywords

Agarista salicifolia, Antibiotics, Assimilation rate, Inhibition zone, Ensete ventricosum,, Induced resistance, Methanol extracts, Phenylalanine ammonia lyase, Total phenolic compounds. Xanthomonas campestris pv. musacearum

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