Unraveling the dynamics of wheat leaf blight complex: isolation, characterization, and insights into pathogen population under Indian conditions

Abstract

Wheat, a staple food crop for 35% of the global population, faces a threat from Helminthosporium leaf blight (HLB), a complex of spot blotch (Bipolaris sorokiniana) and tan spot (Pyrenophora-tritici-repentis) diseases under warm and humid conditions. However, in Indian conditions, the knowledge of existing pathogen populations associated with the HLB complex is limited and largely dominated by only B. sorokiniana (spot blotch). To address this, diseased samples were collected from all six wheat growing zones during 2020-2022. The pathogenic species were identified through in-depth morphological characterization, supplemented with ITS-rDNA and GAPDH sequence analysis, a diagnostic SCAR marker, and pathogenicity studies on two wheat varieties: Sonalika and HD2733. The 32 isolates collected from 10 different states consist of B. spicifera (12.5% of all isolates), Exserohilum rostratum (9.3%), Bipolaris oryzae (3.1%), and B. sorokiniana (75%). B. sorokiniana exhibited the highest disease severity on both varieties. Other lesser-known pathogenic species also produced comparable disease severity as B. sorokiniana isolates and, therefore are economically important. Unraveling pathogen composition and biology aids in disease control and resistance breeding. Our study highlights economically impactful and lesser-known pathogenic species causing wheat leaf blight/spot blotch in India, guiding both current management and future resistance breeding strategies in plant pathology.

Keywords: Bipolaris spicifera; Exserohilum rostratum; absolute quantification; leaf blight/spot blotch complex; soil population dynamics; wheat.

Figure 1

Purified mono-conidial cultures of 32 leaf blight/spot blotch isolates on potato dextrose agar slants (15 days old).

Figure 2

Scatterplot representing the mycelial growth rate of 32 leaf blight/spot blotch isolates on potato dextrose agar medium over a period of 15 days. Dots represent 32 isolates at 3, 6, 9, 12, and 15 days after plating on PDA medium. Size of the dots indicates their average growth rate/day (mm/day). Color of dots represents the two categories of isolates: “Group 1” isolates are slow-growing with a lesser average growth rate (<5 mm/day) and “Group 2” isolates are fast-growing with a higher average growth rate (>5 mm/day).

Figure 3

Phylogenetic analysis of rDNA-ITS region of fungal isolates obtained from leaf blight-infected wheat leaves and seeds in wheat growing zones of India. The evolutionary analysis of Bipolaris sorokiniana, Bipolaris spicifera, Bipolaris oryzae, and Exserohilum rostratum is inferred based on rDNA-ITS sequences using the maximum likelihood method with 1,000 replications. Reference sequences (5 for each cluster) retrieved from NCBI are provided with accession numbers along with species names. The rDNA-ITS sequence of Magnaporthe oryzae was taken as an outgroup.

Figure 4

Phylogenetic analysis of the GAPDH gene of 32 fungal isolates obtained from leaf blight-infected wheat leaves and seeds in wheat growing zones of India. The evolutionary analysis of Bipolaris sorokiniana, Bipolaris spicifera, Bipolaris oryzae, and Exserohilum rostratum is inferred based on GAPDH sequences using the maximum likelihood method with 1,000 replications. Reference sequences along with sequences of ex-type species retrieved from NCBI are provided with accession numbers. The GAPDH sequence of Fusarium oxysporum was taken as an outgroup.

Figure 5

Varietal difference between two wheat varieties, HD2733 (moderately resistant) and Sonalika (susceptible), on the basis of (A) disease severity (in percentage), and (B) lesion size (in cm²) produced by 32 leaf blight/spot blotch isolates taken for the study.

Figure 6

Grouped Barplot representing disease severity, measured as average disease index (ADI), of 32 leaf blight isolates on Sonalika (susceptible variety) and HD2733 (moderately resistant variety). Bars represent standard deviation of the mean.

Figure 7

Box plot of soil population dynamics studies of 32 leaf blight isolates in the rhizospheric zone of (A) Sonalika (susceptible variety), and (B) HD2733 (moderately resistant variety). Through absolute quantification, DNA copy number/g soil was calculated for all isolates at 15-day intervals for a period of 3 months. The lines outside the box extend to the highest and lowest observations. The lower and upper hinges correspond to the first and third quartiles. A line across the box represents the sample median and the small black circles represent outliers.

Figure 8

Pictorial illustration of four pathogenic species identified during the study associated with leaf blight/spot blotch complex under Indian climatic conditions.

Figure 9

Correlogram representing Pearson’s correlation coefficient for disease severity caused by 32 leaf blight/spot blotch isolates, measured as Average Disease Index (ADI) with fungal growth parameters (lesion size, growth rate on PDA medium, sporulation on PDA medium and soil population) on (A) Sonalika (susceptible variety) and (B) HD2733 (moderately resistant variety).