Methods and tools for diagnosis of plant diseases: the case of tip rot of carrot

Tip rot of carrot significantly reduces root quality and contributes to the high-level rejection during sorting and packaging in Norway. The rot can be dry, or wet, and vary in colour from light brown to dark brown. Diagnosis of a plant disease involves close examination of the symptoms, detection and identification of the causal agent(s), and confirmation of pathogenicity. The objective of this study was to identify the causal agent(s) of tip rot in carrot. Fungi and bacteria were isolated from multiple carrots with tip rot symptoms and used for inoculation of healthy carrots to determine pathogenicity and also for DNA extraction, sequencing of commonly used genes for identification and barcoding genes and DNA metabarcoding. For isolation and inoculation, we developed a method allowing individual carrots to remain upright without touching each other within an incubation box. For morphological identification of causal agents, we found that a combination of methods such as isolation on potato carrot agar, disinfection of infected tissue followed by moist incubation, and inoculation followed by incubation at room temperature for 24 h, and then at 0-6°C were optimal methods for the identification of tip rot pathogens of carrot. Based on the combination of molecular and morphological identification methods, we found that tip rot of carrots is a disease complex caused by several fungi but principally Mycocentrospora acerina and Cylindrocarpon destructans. Diagnosis of postharvest diseases is often a complex problem, and this research demonstrates that a combination of methods is a useful approach. Furthermore, the study indicated that the common approach of trying to associate a disease with a single causal agent does not work for all postharvest diseases. The possibility of multiple causal agents and predisposing factors must be considered, and we should be cautious not to jump to a hasty decision.