Identification of Pseudopestalotiopsis ampullacea as a new pathogen causing tea gray blight in India and its management strategies

Abstract

Gray blight is a serious threat to the tea [Camellia sinensis (L.) O. Kuntze] production in major tea cultivating countries including India. The disease is caused by Pestalotiopsis-like species. In this study, five isolates of Pseudopestalotiopsis species isolated from symptomatic tea leaf samples in North Bengal, India were investigated. Based on the multi-locus phylogenetic analysis using concatenated sequences of three (ITS, tef-1 alpha, and tub-2) loci, cultural and micromorphological characters, and host association, the fungal isolates were identified as Pseudopestalotiopsis ampullacea F. Liu & L. Cai. The morphological analysis also revealed that the fungal isolates were evidently differentiated from other Pseudopestalotiopsis species. To date, P. ampullacea has not been reported on tea plants in India. Among the five isolates studied, isolate NKT0P03 was randomly selected for pathogenicity tests and its sensitivity to fungicides and microbial antagonists. In pathogenicity test, the isolate showed weak to high virulence reactions on 25 different tea cultivars. The pathogen showed an avirulent reaction on the cultivar TV11. In order to identify an effective management strategy against this new pathogen, synthetic fungicides and microbial biocontrol agents were evaluated in the laboratory. Results revealed that carbendazim + mancozeb, hexaconazole, propiconazole, and valextra were effective fungicides with an 85.1% to 89.8% range of inhibitory activity against P. ampullacea NKT0P03. Among microbial agents, Trichoderma harzianum, T. reesei, T. hamatum, Bacillus subtilis, and Microbacterium barkeri were efficient bioagents against P. ampullacea NKT0P03 with antagonistic activity ranging between 66.6% and 84.2%. Thus, these fungicides and microbial bioagents can be recommended as effective agents for the management of P. ampullacea causing tea gray blight after their field evaluations.

Keywords: Microbial agents; Pathogenicity; Phylogeny; Synthetic fungicides.

Fig. 1

Symptoms developed by Pseudopestalotiopsis ampullacea and its cultural characteristics on agar plate, images (a) and (b) are showing initial necrotic lesions developed by the pathogen, (c) is advanced symptom, and its conidiomata (d) obverse (e) and reverse (f) colony morphology of representative isolate NKT0P03 on agar plate.

Fig. 2

Tea garden (Nagrakata) showing leaf defoliation caused by gray blight (left; arrow indicates defoliated plants) and a tea plant infected with gray blight (right; arrow indicates gray blight symptomatic leaves).

Fig. 3

Conidial morphology of P. ampullacea isolates NKT0P03 (a, b), NKT0P04 (c), NKT0P05 (d), NKT0P06 (e), and NKT0P07 (f) on PDA medium after 8 days incubation (scale bar = 10 µm).

Fig. 4

Phylogenetic tree obtained using the MP method based on a combined datasets of ITS, tef-1 alpha, and tub-2 genes sequences of 25 strains showing the relationship of isolates NKT0P03, NKT0P04, NKT0P05, NKT0P06 & NKT0P07 with Pseudopestalotiopsis species, and with the Neopestalotiopsis species as the outgroup taxon. The numbers in the bootstrap test (1000 replicates) are indicated above and below the branches. Bootstrap support values for MP greater than 50% are shown left at the nodes and Bayesian posterior probabilities greater than or equal to 0.95 are indicated right at the nodes. Scale bar indicates expected number of changes per site.

Fig. 5

Typical symptoms developed by representative isolate P. ampullacea NKT0P03 on the leaves of various tea cultivars, cultivar TV11 is showing immune reaction.

Fig. 6

Heat map showing relationship between tea cultivars, lesion diameter (LD), and percent disease incidence (DI). The heatmap was generated using online software http://heatmapper.ca/.