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Wood rot fungus <i>Fulvifomes siamensis</i> infects multiple urban tree species commonly planted in Singapore. A commercial e-nose (Cyranose 320) was used to differentiate some plant and fungi volatiles. The e-nose distinctly clustered the volatiles at 0.25 ppm, and this sensitivity was further increased to 0.05 ppm with the use of nitrogen gas to purge the system and set up the baseline. Nitrogen gas baseline resulted in a higher magnitude of sensor responses and a higher number of responsive sensors. The specificity of the e-nose for <i>F. siamensis</i> was demonstrated by distinctive clustering of its pure culture, fruiting bodies collected from different tree species, and in diseased tissues infected by <i>F. siamensis</i> with a 15-min incubation time. This good specificity was supported by the unique volatile profiles revealed by SPME GC-MS analysis, which also identified the signature volatile for <i>F. siamensis</i>—1,2,4,5-tetrachloro-3,6-dimethoxybenzene. In field conditions, the e-nose successfully identified <i>F. siamensis</i> fruiting bodies on different tree species. The findings of concentration-based clustering and host-tree-specific volatile profiles for fruiting bodies provide further insights into the complexity of volatile-based diagnosis that should be taken into consideration for future studies.