Morpho-cultural and molecular variability of Stemphylium vesicarium causing Stemphylium leaf blight in tropical...

The cultural, morphological, and molecular variations among 22 isolates of Stemphylium vesicarium (Wallr.) E. Simmons, collected from different locations, was carried out. Significant variations were observed among the Stemphylium isolates regarding colony diameter, mean radial growth rate and sporulation on PDA media. The colonies of S. vesicarium isolates exhibited diverse mycelial growth characteristics, including velvety, cottony, or fluffy textures and a range of colours from whitish to dark grey, olivaceous with a greenish tinge or brownish. The margins of the colonies were observed to be filiform, displaying a filamentous appearance, with entire and undulate shapes and a whitish colouration. The conidiophores of Stemphylium isolates displayed a wide range of dimensions, with average length ranging from 36.81 μm to 66.44 μm and average breadth from 3.05 μm to 6.96 μm. Similarly, the conidia exhibited variations in size, colour, shape, average length (23.31 μm-43.18 μm) and average breadth (12.84 μm-23.13 μm). The conidia were mainly light brown or brown and displayed ovoid, oblong, or ovoid-to-oblong shapes. The number and presence of transverse and longitudinal septa also varied among the isolates. Fifteen RAPD primers generated 192 banding patterns. SV4, OPL5, and SV5 were identified as the most polymorphic primers, while OPA3, OPF10, OPN7, OPS7, and OPS10 produced the fewest polymorphic bands. The average polymorphic information content (PIC) value was 0.37, with OPA5 and OPC8 showing the highest PIC values. Cluster analysis based on genetic similarity revealed five distinct clusters, but no clear correlation between isolates and their collection sites was observed. In the phylogenetic analysis, based on Internal transcribed spacer (ITS) region and Glycerol-3-Phosphate Dehydrogenase 1(gpd1) gene sequences, 20 isolates obtained from diseased onion leaves formed a distinct cluster and exhibited sequence similarity with ex-type sequence of Stemphylium vesicarium. Additionally, two isolates from diseased garlic samples showed similarity with ex-type sequence of Stemphylium eturmiunum. This is the first-time report of S. eturmiunum on Indian garlic under field conditions.