Management of Charcoal Rot (Macrophomina phaseolina) of Fenugreek in Rajasthan

Fenugreek (Trigonella foenum-graecum L.) also known as Methi belongs to family leguminoceae. Fenugreek believed to be originated from South East Europe and West Asia is cultivated throughout India and other parts of world for leafy vegetables, condiment, medicinal purposes and fodder also. The leaves (per 100 g of edible portions) contain moisture (per 100 g of edible portions) 86.1 g, thiamine 0.05 mg, fat 0.9 g, calcium 360 mg, protein 4.4 g, oxalic acid 13 mg, fiber 1.1 g, iron 17.2 mg, potassium 51 mg, mineral 1.5 g, sulphur 167 mg, carbohydrates 6.00 g, vitamin A 6450 IU, magnesium 67 mg, nicotinic acid 0.7 mg, sodium 76.1 mg, vitamin C 54 mg, phosphorus 51 mg and chlorine 165 mg (Bose and Som, 1986).
       
India stands first in the production of fenugreek with total acreage of 1.20 lakh  hectares and production of 1.88 lakh tonnes with average productivity of 1566.23 kg/ha. In India more than 80 per cent area and production of is contributed by Rajasthan state alone. In Rajasthan first rank in the production of fenugreek with total acreage of 45.31 thousand hectares and production of 62.89 thousand tonnes with average productivity of 1388.00 kg/ha (Anonymous, 2019-20).
       
Fenugreek is cultivated in almost all districts of Rajasthan whereas Bikaner district stands first in production with total acreage of 7046 hectares and production of 7760 tonnes with productivity of 1101 kg/ha (Anonymous, 2018-19).
       
Fenugreek is infected by several fungal, bacterial and viral diseases. Major fungal diseases of fenugreek are cercospora leaf spot (Cercospora traversian), charcoal rot (Macrophomina phaseolina (Tassi) Goid.), wilt (Fusarium oxysporum Schlecht.), downy mildew (Peronospora trigonellae Gaum), rhizoctonia root rot (Thanatephorus cucumeris Kuhn), powdery mildew (Leveillula taurica (Lev.) Arm), root rot (Sclerotinia trifoliorum Sacc.) and rust (Uromyces trigonellae Pass.) etc.
       
Among all the fungal diseases of fenugreek, the most destructive is charcoal rot of fenugreek caused by M.  phaseolina causing potential yield losses. Charcoal rot infected seeds act as an important source of primary inoculums for new areas. Plant stand affected due to pre and post emergence infection of crop. In pre emergence stage, the fungus causes seed rot and mortality of germinating seedling while in post emergence stage seedling get blightened due to soil and seed borne infection. The pathogen may infect almost all parts of plants viz., root, stem, branches, petioles, leaves and pods.

Efficacy of fungicides against M. phaseolina in vitro
 
The efficacy of fungicides and combination products were tested by using 100, 200, 300, 400 and 500 ppm against M. phaseolina in vitro. Eight fungicides were evaluated in this study. These were T₁: captan 70% + hexacunazole 5% WP, T₂: azoxystrobin 18.2% + difenaconazole 11.4% SC, T₃: chlorothalonil 75% WP, T₄: carbendazim 12% + mancozeb 63% WP, T₅: tebuconazole 50% + trifloxystrobin 25% WG, T₆: carboxin 37.5% + thiram 37.5% WP, T₇: copper oxy chloride 50% WP and T₈: captan 70% WP.
       
The efficacy against mycelial growth was tested using Poisoned-Food-Technique (Nene and Thapliyal, 1973). Required quantities of fungicide was thoroughly mixed in melted PDA, just before pouring in sterilized petri dishes and were allowed to solidify for 12 hrs. Each plate was then inoculated with 5 mm disc of mycelial bit taken from the periphery of 10 days colony of M. phaseolina growing on PDA. The inoculated petri dishes were incubated at 25±1°C. Three plates were used for each treatments. serving as three replications. Colony diameter (two diagonals) was measured after 15 days of incubation. Medium without fungicide served as control. Per cent growth inhibition was calculated by Vincent’s (1947) formula as follows:
   
Where,
C= Mycelial growth of M. phaseolina in control (average of both diagonals).
T= Mycelial growth of M. phaseolina in treatment (average of both diagonals).
 
Efficacy of bioagents and fungicides against M. phaseolina in vivo
 
A field trial was conducted for the management of charcoal rot of fenugreek variety Rmt-305 using bio-agents and fungicides in Rabi season 2020-21. A most popular fenugreek cultivar Rmt-305 was used in this experiment. The fenugreek was sown on 29^th October 2020. The crop was planted at 30 cm row to row and 10 cm plant to plant spacing. The gross plot size was 3.0 × 3.0 m². The experiment was laid out in randomized block design (RBD) with three replications. All other recommended practices required for cultivation of the crop were followed. Talc based formulations of one fungal and one bacterial bio-agent viz., T. harzianum, P. fluorescens were applied as seed treatment and soil application @ 5 + 5 g/kg and @10 kg/ha respectively and fungicide tebuconazole 50% + trifloxystrobin 25% @ 1.5 g/kg, carbendazim 12% + mancozeb 63% @ 2 g/kg and azoxystrobin 18.2% + difenaconazole 11.4% @ 2 ml/kg seed were used. Seed treatment of fungicides, seed and soil treatment of the talc based bio-agent formulations was done. In case of control, seeds were sown in pathogen inoculated soil without any bio-agents and fungicides. The combination of the treatments and their doses were mentioned in Table 1. Observations on charcoal rot incidence were recorded periodically as well as the grain yield was recorded after harvesting of the crop.
 

 
Calculation and statistical analysis
   
The data of per cent disease incidence in all the experiments were transformed to their Arc sin values (Fisher and Yates, 1963). The statistical analysis of the data of all the laboratory experiments was done following completely randomized design. The data of field Experiments were analysed following randomized block design (Cochran and Cox, 1957).

Efficacy of fungicides against M. phaseolina in vitro
 
Eight (systemic, contact and combi) fungicides were tested at different concentration of 100, 200, 300, 400 and 500 ppm against M. phaseolina using Poisoned Food Technique in vitro. Table 2 and Plate 1 shows the effectiveness of all the tested fungicides. The data showed that the maximum mycelial growth inhibition was increased at different concentration of fungicides. tebuconazole 50% + trifloxystrobin 25% WG was found Cent percent inhibition of mycelial growth of M. phaseolina  followed by captan 70% + hexaconazole 5%WP (2.93 mm) and carbendazim 12% + mancozeb 63% WP (10.26 mm) resulting in significantly reduction of mycelial growth of M. phaseolina. At 200 ppm concentration, growth of mycelium was not observed in tebuconazole 50% + trifloxystrobin 25% WG (0.00 mm) and captan 70%+ hexaconazole 5% WP (0.00 mm) followed by carbendazim 12% + mancozeb 63% WP (5.84 mm). At 300 ppm concentration, tebuconazole 50% + trifloxystrobin 25% WG (0.00 mm), captan 70% + hexaconazole 5%WP (0.00 mm) and carbendazim 12% + mancozeb 63% WP (0.00 mm) inhibited Cent per cent mycelium growth. At 400 ppm and 500 ppm concentration, no mycelial growth of fungus was recorded in tebuconazole 50% + trifloxystrobin 25% WG, captan 70%+ hexaconazole 5%WP and carbendazim 12% + mancozeb 63 % WP (0.00 mm).
 

 

       
Similarly, treatment Tebuconazole 50% + trifloxystrobin 25% WG was most effective at lowest concentrations viz., 100, 200 and 300ppm. Similar, results was observed by Kumar et al., (2016) this study suggests the possibility to enhance the antifungal activity of fungicides Tebuconazole 50% + trifloxystrobin 25% WG towards the control of M. phaseolina. Bashir et al., (2017) The interaction between treatments and concentrations (T×C) showed that used concentrations 150 ppm, 250 ppm and 350 ppm of Nativo abundantly inhibit fungal colony growth up to 1.26 cm, 0.86 cm and 0.66 cm, respectively whereas the interaction between treatments and days expressed that after day ninth the minimum colony growth (1.23 cm) was observed for Nativo as compared to all other treatments.
 
Efficacy of bioagents and fungicides against M. phaseolina in vivo
 
The results showed in Table 3 indicate that the charcoal rot severity in fenugreek was effectively reduced by combination of bioagents and fungicides viz., seed treatment with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + soil application of T. harzianum @ 10 kg/ha + soil drenching with tebuconazole 50% + trifloxystrobin 25% WG  (6.94%) followed by ST with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + SA of P. fluorescens @ 10 kg/ha + SD with tebuconazole 50% + trifloxystrobin 25% WG (9.76%) and ST with carbendazim 12% + mancozeb 63% WP @ 2 g/kg seed + SA of T. harzianum @ 10 kg/ha + SD with carbendazim 12% + mancozeb 63% WP (13.22%) as compared to control where highest disease severity of 48.55% was recorded. Among the treatments highest disease severity was recorded in bioagents ST @ 10 g/kg seed with P. fluorescens + SA of P. fluorescens + SD of P. fluorescens (24.35%) followed ST with T. harzianum + SA of T. harzianum + SD of T. harzianum (21.22%) and ST with azoxystrobin 18.2% + difenaconazole 11.4% SC @ 2 ml/kg seed (18.31%).
       

 
The maximum disease control was recorded in seed treatment with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + soil application of T. harzianum @ 10 kg/ha + soil drenching with tebuconazole 50% + trifloxystrobin 25% WG (85.72%) followed by ST with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + SA of P. fluorescens @ 10 kg/ha + SD with tebuconazole 50% + trifloxystrobin 25% WG (79.85%) and ST with carbendazim 12% + mancozeb 63% WP @ 2g/kg seed + SA of T. harzianum @ 10 kg/ha + SD with carbendazim 12% + mancozeb 63% WP (72.77).  Disease incidence of charcoal rot in fenugreek under field conditions and a view of experimental field are shown in Plate 2.
 

 
Grain yield
 
Maximum grain yield of fenugreek (19.83 q ha^-1) was obtained in seed treatment with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + ST @ 10 kg/ha of T.  harzianum @ 10kg/ha + SD @ 2g/L with tebuconazole 50% + trifloxystrobin 25% WG, followed by ST with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + SA @ 10 kg/ha of P. fluorescens @ 10 kg/ha + soil drenching @ 2 g/L with tebuconazole 50% + trifloxystrobin 25% WG (18.60 qha^-1) and ST with carbendazim 12% + mancozeb 63% WP @ 2 g/kg seed + SA of T. harzianum @ 10 kg/ha + SD with carbendazim 12% + mancozeb 63% WP (17.20 qha^-1). Similar findings was recorded by Meena et al., (2018) efficacy of Tebuconazole 50% + trifloxystrobin 25% WG, Carbendazim 12% + mancozeb 63% WP, T. harzianum and P. fluorescens were assessed under in vivo conditions. Bashir et al., (2017) observed that in the field conditions the treatment Nativo was found minimum disease incidence. These findings are very much similar with our findings.

It can be concluded unequivocally considering the results that in field conditions combination of bioagents and fungicide viz., seed treatment with tebuconazole 50% + trifloxystrobin 25% WG @ 1.5 g/kg seed + SA of T. harzianum @ 10 kg/ha + SD @ 2 g/L with tebuconazole 50% + trifloxystrobin 25% WG was found most effective in managing the charcoal rot disease, lowest disease severity and maximum grain yield.

The authors are very much thankful to Head Department of Plant Pathology, College of Agriculture, Swami Keshwanand Rajasthan Agricultural University, Bikaner for providing facilities to conduct the experiment.

None.