Effect of bioinoculants on rhizome rot disease of turmeric (Curcuma longa L.)

Introduction

The world's largest producer, consumer and exporter of spices is India, often referred as the land of spices. Cardamom and black pepper are the two commodities that yield the most foreign exchange, followed by turmeric (Curcuma longa L.). About 80 percent of global production and 60 percent of exports are attributed to India, which is the leading producer of turmeric worldwide. 5,34,000 tonnes of turmeric are produced annually on 1,34,000 hectares of land in India (Ravikumar, 2002) [15] . Diseases such as rhizome rot, anthracnose and leaf blight affects turmeric. Ramarethinam and Rajagopal (1999) [13] indicated that rhizome rot incited by Pythium sp. is a major constraint in all areas of India that grow turmeric. According to Rathiah (1987) [14] , the disease caused up to 95% of crop yield loss due to symptoms such as collapse of tillers, rotting roots and the hollow rhizome leaving only fibrous tissues. According to Nirmal (1992) [9] , the disease results in 50-80% yield loss during storage and more than 60% seedling mortality in both nursery and field conditions. Rhizome rot also causes a 50% yield loss (Rajalakshmi et al., 2016) [12] . Rhizome rot has become more common as a result of intensive turmeric farming. Fungicide-induced rhizome rot control has decreased soil microbial biodiversity and led to the emergence of pathogen strains that are resistant to the disease. The need for alternate plant protection techniques has arisen from growing concerns about the effects of pesticide misuse on the environment. The present research thus focuses into the use of antagonistic bioinoculants to manage turmeric rhizome rot in a way that's environmentally friendly.

Under glass house conditions, P. aphanidermatum multiplied on sand maize medium was added at a rate of 5% (w/w). Data on the percentage of rhizome rot occurrence was recorded during harvest. Additional growth characteristics such as plant height, leaf count and stem girth were also recorded. Eighteen treatments were used in the experiment under CRD design which was replicated three times. The particulars of each treatment include: T 1 -P. chlororaphis PA23 (RD) ; T 2 -P. chlororaphis PA23 (RD) + SA on 3 rd Month T 3 -P. chlororaphis PA23 (RD) + SA on 5 th Month T 4 -P. chlororaphis PA23 (RD) + SA on 3 rd and 5

Effect of bioinoculants on defense-related enzyme induction in turmeric plants

The bioinoculants viz., B. subtilis CBE4, P. chlororaphis PA23 and T. viride MNT7 were applied to three-month-old turmeric plants through soil and they were then challenged with P. aphanidermatum. After six days of challenge inoculation with P. aphanidermatum, the turmeric leaves were carefully removed from the pots and repeatedly rinsed with distilled water in order to extract enzymes. The green leaves were chilled at 5 °C and homogenized using 0.1 M phosphate buffer (pH 7.0) and further centrifuged at 5 °C for extracting the enzymes. The supernatant served as source for analysis of defense related enzyme for colorimetric estimation of chitinase (Boller and Mauch, 1988) [2] , -1, 3glucanase (Pan et al., 1991) [10] , peroxidase (Hammerschmidt et al., 1982) [6] , poly phenol oxidase (Mayer et al., 1965) [7] . phenyl alanine ammonia lyase (Dickerson et al., 1984) [5] , phenols (Zieslin and Ben Zaken, 1993) [20] and protein (Bradford 1976) [4] . The data were statistically analyzed using the OP STAT software.

Results and Discussion

The effectiveness of the bioinoculants in managing turmeric rhizome rot was assessed in glass house system. The outcomes are summarized in Table 1. Delivering the bioformulation P. chlororaphis PA23 and B. subtilis CBE4 as rhizome dip and soil treatment on 3 rd and 5 th month (T 12 ) exhibited the lowest disease incidence (14.60 percent). Rhizome dip and soil treatment on the third and fifth months of planting using B. subtilis CBE4 (T8) and P. chlororaphis PA23 (T4) resulted in disease incidence of 20.00 and 20.83 percent respectively which are statistically comparable. The maximum disease incidence of 52.66 percent was observed in the control plants. The results align with the findings of Muthulakshmi and Saveetha (2009) [8] , that introducing consortia of Trichoderma viride and Pseudomonas flourescens by seed and soil application was an efficient way to reduce the severity of turmeric rhizome rot disease with maximizing yield. In both glasshouse and field conditions, the rhizome rot disease incidence was lowest when Pseudomonas fluorescens FP7 liquid formulation was used as rhizome dip and soil drench ( Prabhukarthikeyan et al., 2017) [11] . Ginger rhizome rot can be effectively managed by applying T. harzianum as rhizome dip and soil at 30, 60, and 90 days post-planting (Shanmugam et al., 1999) [17] . According to Ramarethinam and Rajagopal (1999) [13] , rhizome rot of turmeric was suppressed by applying Trichoderma. Application of bioinoculants significantly increased the yield, plant height, stem girth and no. of leaves/ clump of turmeric plants. Treatment [19,1] . Instead of applying bioinoculants individually through rhizome dip or soil treatment on 3 rd and 5 th Month, application of bioinoculants as a consortial formulation was effective. Due to the pathogen's mobility and potential to reinfect seed at a later stage of treatment, soil application of bioinoculants was shown to be more successful than treating rhizomes. However, the application of bioinoculants in soil led to a significant increase in the antagonistic population, which significantly slowed the growth of the pathogen. This could be the cause for the decreased occurrence of rhizome rot in treated plots.

Impact of bioinoculants on induction of defense related enzyme in turmeric plants

One novel method of protecting plants is to use biological stimulants to activate the plant's defense mechanisms. Table 2 presents the findings of the induction of defense enzymes, phenol and protein following the challenge inoculation of P. aphanidermatum in soil after bioinoculants treatment. When exposed to pathogens in plants treated with bioinoculants, the turmeric plants exhibited increased chitinase activity. The pathogen's challenge inoculation significantly increased the activity. When P. chlororaphis PA23 + B. subtilis CBE4 were applied to turmeric plants followed by challenge inoculation with P. aphanidermatum, the plants' chitinase activity was increased (400 n mol). In the healthy control, chitinase activity (250.00 n mol of) which remained at low level. Turmeric plants' resistance to P. aphanidermatum infection was considerably enhanced by a significant rise in chitinase activity (Sathiyanarayanan and Muthukrishnan, 2014) [16] Following the application of bioinoculants, the turmeric plants exhibited increased -1, 3, glucanase activity in response to pathogen stimulation. The healthy control group showed reduced -1,3 glucanase activity with a level of 112.15 mol. When P. chlororaphis PA23 and B. subtilis CBE4 were applied together followed by a challenge infection with P. aphanidermatum, the activity of -1, 3 glucanase was increased (322.58 mol). P. aphanidermatum inoculated plants that were pretreated with bioinoculants showed a greater concentration of this hydrolytic enzyme. When the plants were primed with bioinoculants and challenged with pathogen the turmeric plants showed increased PO activity. Plants treated with bioinoculants alone also showed increased peroxidase activity. However, plants that had been pretreated with bioinoculants and P. aphanidermatum challenge inoculation showed an additional rise in activity. The findings showed that turmeric plants pretreated with P. chlororaphis PA23 + B. subtilis CBE4 and challenge inoculation with P. aphanidermatum had a considerably higher induction of PO enzyme (4.083). However, the healthy control plants had very less PO activity (1.520). When exposed to a pathogen, the turmeric plants shown increased PPO activity irrespective of the fact they were previously treated with bioinoculants. Compared to the uninoculated control, the aforesaid treatment resulted in a two-fold higher induction of PPO. P. chlororaphis PA23 + B. subtilis CBE4 pretreatment of turmeric plants, followed by P. aphanidermatum challenge inoculation, resulted in the highest PPO activity (2.317). However, in the healthy control, the activity was incredibly low (1.117). Nevertheless, the activity in bioinoculants pretreated plants that were not challenged with a pathogen did not significantly decrease. When exposed to a pathogen, the turmeric plants had increased PAL activity regardless of whether they had previously been treated with bioinoculants. The highest PAL activity (12.325  mol) was reported by pretreating turmeric plants with P. chlororaphis PA23 + B. subtilis CBE4 followed by challenging with P. aphanidermatum. The activity of PAL in the pathogen-inoculated control was 8.513  mol, while the activity of PAL was 6.233  mol in the healthy control. Following challenge inoculation with pathogen, phenolics accumulation increased. The total phenol content of fresh tissue treated with P. chlororaphis PA23 + B. subtilis CBE4 was 600 g g -1 following pathogen inoculation. However, pathogen-inoculated control the total phenol was only 420 g g -1 . It was evident that, in comparison to plants treated with bioinoculants and challenged with pathogen, the accumulation of phenolic compounds was relatively lower in the healthy control (350 g g -1 ). When P. aphanidermatum was applied to turmeric plants pretreated with bioinoculants, the plants showed increased protein accumulation. When compared to the other treatments, the activity was much higher (490 g/g) in turmeric plants primed with P. chlororaphis PA23 + B. subtilis CBE4 and challenge inoculated with P. aphanidermatum. In contrast, healthy control recorded protein accumulation of 290 g/g, the pathogen-inoculated control reported protein accumulation of 355 g/g. Pretreatment of turmeric plants with bioinoculants followed by challenging with P. aphanidermatum, induced higher accumulation of protein. The activity was significantly higher in leaves (490 g/g) of turmeric plants pretreated with chlororaphis PA23 + B. subtilis CBE4 treatment challenge inoculated with P. aphanidermatum when compared to all other treatments. The pathogen inoculated control recorded protein accumulation (355 g/g) when compared to the healthy control which recorded protein accumulation of 290 g/g. Several plant defense enzymes were activated when FYM-based biomanure of T. viride was applied to turmeric plants ( Ushamalini et al., 2006) [18] . The results are similar to those of Vinayarani et al., 2019 [19] , that defense enzymes and phenolics were increased by two to three times when T. asperellum was pretreated in turmeric followed by challenge inoculation with pathogen when compared to the untreated control.

Conclusion

The results of the present investigation revealed that application of bioinoculants B. subtilis CBE4 and P. chlororaphis PA23 as rhizome and soil application in turmeric suppressed rhizome rot and enhanced plant growth and induced the defense enzymes.

References

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