I. R. M.
A. C. R. C.
Scientia Terrae Research Institute, Fortsesteenweg 30A, B-2860 Sint-Katelijne-Waver, Belgium
B. P. A.
Centre of Microbial and Plant Genetics (CMPG), Katholieke Universiteit Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium
B. P. H. J.
Wageningen University, P.O. Box 8025, 6700 EE Wageningen, the Netherlands
In January 2003, a severe root and foot rot was observed on 2-month-old wilted tomato (Lycopersicon esculentum Mill.) plants in a large-scale (2.5 ha) commercial greenhouse setting in Belgium. Tomato plants (10%) produced from healthy nursery-grown seedlings and planted to new, clean rockwool and drip irrigation with UV-disinfected water developed symptoms. Symptom development was restricted to lower plant parts with severe rotting of the entire root system and dark lesions girdling the stem base. No symptoms of disease were observed on the foliage or upper stems. Cross sections of the stem base revealed brown discoloration of internal tissue, including the vascular tissue and pith. Dark brown lesions also occurred on the roots. Sections of the stem base, the upper roots (sampled near to the stem base), and the lower roots (sampled on roots deeper in the rockwool) were plated separately on corn meal agar. The oomycete pathogen Phytophthora infestans (Mont.) de Bary was identified in each sample on the basis of morphological characteristics observed directly with light microscopy. Branched sporangiophores with slight swellings and characteristic lemon-shaped sporangia (35 × 20 μm and ratio length/width of 1.75 μm) at their tips were obvious after incubation in darkness at 24°C. Oospores and chlamydospores were not observed. After multiple soil treatment with oomycete-specific fungicides, the plants recovered. Since the occurrence of P. infestans on roots is unusual, the identity of the pathogen on the diseased plant tissues was confirmed with three techniques, DNA array identification, internal transcribed spacer (ITS) sequencing, and a polymerase chain reaction (PCR) amplification using P. infestans-specific primers. DNA was directly processed from separate samples of upper and lower root and stem base tissue. The DNA array used was originally developed to detect and identify the key fungal pathogens of tomato (2). Among detector probes for other tomato pathogens, this array contains oligonucleotide detector probes for P. infestans (PIN1: 5′-GGT TGT GGA CGC TGC TAT T and PIN2: 5′-AAT GGA GAA ATG CTC GAT TC). These probes are based on ITS sequences (ITS I and ITS II). Using conserved ribosomal primers OOMUP18Sc (5′-TGC GGA AGG ATC ATT ACC ACA C) and ITS4, oomycete DNA was amplified by PCR and simultaneously labeled with alkaline-labile digoxigenin (2). All generated amplicons strongly hybridized to the oligonucleotide detector probes for P. infestans and not to any other pathogen-specific detector probe present on the array. The pathogen could not be detected in roots and stem bases of symptomless plants. In addition, the ITS-region was sequenced and showed 100% homology with multiple GenBank accessions of P. infestans sequences. As a third confirmatory test, a PCR was performed on DNA extracts from infected root and stem base tissues using a primer set specific to P. infestans (O8-3/O8-4 ). A band of the expected size was produced for the infected stem base and root samples. Until now, this pathogen was known worldwide to cause late blight on potatoes and tomatoes. To our knowledge, this is the first report of root and foot rot of tomato caused by P. infestans.
References: (1) H. S. Judelson and P. W. Tooley. Phytopathology 90:1112, 2000. (2) B. Lievens et al. FEMS Microbiol. Lett. 223:113, 2003.