Screening and Field Validation of Fungicides for Control of Alternaria Spot in Pomegranate (Punica granatum L.)

Introduction

Pomegranate (Punica granatum L.) is an important fruit crop cultivated in arid and semi-arid regions due to its adaptability to adverse environmental conditions (Jalikop, 2010; Levin, 2006) [8,10] . In India, it has gained significant commercial importance, with Maharashtra being the leading producer (Anonymous, 2023) [2] . The fruit is valued for its nutritional and medicinal properties, as it is rich in antioxidants, vitamins, and bioactive compounds (Viuda-Martos et al., 2010) [13] . However, pomegranate production is severely affected by various diseases, among which Alternaria leaf and fruit spot caused by Alternaria alternata has emerged as a major threat (Gade and Ghewande, 2010) [7] . The disease leads to significant yield losses and deterioration of fruit quality, especially under warm and humid conditions (Archana and Jamadar, 2014) [3] . Symptoms include dark, concentric lesions on leaves and fruits, resulting in defoliation, fruit rot, and reduced market value (Tziros et al., 2008) [12] . Although fungicides are widely used for disease control, concerns regarding resistance development, environmental safety, and cost have increased (Adesh Kumar et al., 2017) [1] . Therefore, a comprehensive study on the pathogen, its epidemiology, and effective management strategies is essential. The present investigation was undertaken to address these aspects and to develop sustainable disease management practices for pomegranate.

Isolation and Purification of Pathogen (Alternaria alternata).

The pathogen was isolated from naturally infected pomegranate leaves and fruits showing typical symptoms of Alternaria spot (Plate 3,4). The infected plant parts were first washed thoroughly under running tap water to remove surface contaminants and then air-dried. Small tissue bits (2-5 mm) were cut from the junction of healthy and diseased portions using a sterile scalpel. The excised tissue pieces were surface sterilized with 0.1 per cent mercuric chloride solution for one minute, followed by three successive washings with sterile distilled water to remove traces of the sterilant. The sterilized tissues were then blot-dried on sterile filter paper and aseptically transferred onto potato dextrose agar (PDA) medium in Petri plates. The inoculated plates were incubated at 27 ± 2 °C in an inverted position. Fungal growth was observed within 24 hours of incubation, initially appearing white and later turning grey to International Journal of Agriculture and Food Science https://www.agriculturaljournals.com black. The emerging colonies were sub-cultured on fresh PDA medium to obtain pure cultures of the pathogen, which were maintained on PDA slants for further studies.

Morphology of Alternaria alternata

The morphological characteristics of the pathogen were studied on potato dextrose agar (PDA) medium. The colony initially appeared whitish, which later turned grey to black due to profuse sporulation. The growth was cottony to velvety in texture with irregular margins, and the reverse side of the colony exhibited dark pigmentation (Plate 1). Microscopic examination revealed that the mycelium was septate, branched, and light to dark brown in color.

Conidiophores were short, septate, and borne singly or in small groups, measuring approximately 20-60 µm in length and 3-6 µm in width. They were pale to dark brown and slightly swollen at the tip. The conidia were dark brown, produced in chains, and exhibited characteristic obclavate to ellipsoidal shape. They measured about 20-60 µm in length and 7-18 µm in width, with a beak length of 2-10 µm. The conidia were multicellular with 3-10 transverse and 1-5 longitudinal septa (muriform) and possessed a distinct beak (Plate 2). These morphological features confirmed the pathogen as Alternaria alternata.

Pathogenicity Test

Pathogenicity of the isolated fungus was confirmed by inoculating a spore suspension of Alternaria alternata on healthy pomegranate leaves/fruits under humid conditions, while control samples were maintained with sterile water. Typical symptoms of Alternaria spot, such as dark necrotic lesions with concentric rings, developed on inoculated samples (Plate 5), whereas no symptoms appeared in control (Plate 6). The pathogen was successfully re-isolated and found identical to the original culture, thus fulfilling Koch's postulates.

Efficacy of Fungicides against Alternaria alternata under In vitro Conditions

The in vitro evaluation of fungicides against Alternaria alternata revealed significant differences in their efficacy, with mycelial growth inhibition ranging from 66.44 to 90. 1). These findings are in agreement with earlier reports by Dambrauskiene (2006) [4] , Kaur et al. (2020) [9] , and Adesh Kumar et al. (2017) [1] , who also reported high efficacy of azoxystrobin and its combinations against Alternaria spp. under in vitro conditions. After the third spray, disease intensity varied from 5.07 to 37.87 per cent. Azoxystrobin (5.07%) consistently recorded the lowest disease intensity, followed by trifloxystrobin + tebuconazole (5.60%) and meritam + pyraclostrobin (9.07%). The untreated control showed maximum disease intensity (37.87%), while propineb remained least effective (13.87%). These results (Table 2) are in agreement with earlier findings reported by Pasche et al. (2004) [11] , Wang et al.

(2016) [14] , Fagodiya et al. (2022) [6] , and Chakrabarty et al.

(2025) [4] , who also observed superior efficacy of azoxystrobin and its combinations against Alternaria spp.

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Conclusion

The present investigation confirmed Alternaria alternata as the causal agent of leaf and fruit spot of pomegranate based on cultural, morphological, and pathogenicity studies. The pathogen produced characteristic symptoms upon artificial inoculation, thereby fulfilling Koch's postulates.

In vitro evaluation of fungicides revealed that azoxystrobin 23% SC (0.1%) was the most effective in inhibiting mycelial growth, followed by trifloxystrobin + tebuconazole and meritam + pyraclostrobin. Field studies further substantiated these findings, wherein azoxystrobin recorded the lowest disease intensity and highest fruit yield, and was statistically at par with trifloxystrobin + tebuconazole.

Overall, the study demonstrates that effective management of Alternaria spot of pomegranate can be achieved through the application of azoxystrobin or its combination with triazole fungicides. Adoption of these fungicidal treatments can significantly reduce disease severity and enhance yield under field conditions.

References

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