First report of Pseudomonas syringae pv. aesculi on horse chestnut in Switzerland

In 2011 in a public park in Rapperswil-Jona (Switzerland), bleeding cankers (Fig. 1) were observed on 42 out of 300 horse chestnut trees (Aesculus hippocastanum). On March 2015, four to six bark samples were taken from each of three affected trees using a sterilised cork borer (2 mm in diameter). The bark samples were removed from either the centre of the bleeding cankers or from the edge of the necrotic orange-brown lesions (Fig. 2), which became visible after removing the outer bark. Bacterial isolation from the bark samples was made on selective media as described by Green et al. (2009). To detect fluorescent cultures, bacteria were streaked on King’s medium B (King et al., 1954) amended with 50 mg/l cycloheximide. For species identification, the gyrB gene was partially sequenced using the primers gyrB-F/gyrB-R (Green et al., 2009) and the 16S rDNA gene using the primers EUB f933/EUB r1387 (Ji et al., 2004). Six fluorescent bacteria were isolated from all types of samples and from all trees, and were identified as P. syringae pv. aesculi based on 100% identity with 616 bp of the gyrB gene and as Pseudomonas sp. based on 100% identity with 415 bp of the 16S rDNA gene. Two additional non-fluorescent Pseudomonas sp. and Erwinia billingiae were isolated. The Genbank accession numbers of these sequences are KT965182-KT965197.

Pathogenicity tests were performed with two P. syringae pv. aesculi isolates collected during this study (M8597 and M8603), using three-year-old A. hippocastanum saplings. Five replicate saplings were each inoculated through stem cuts with a 5 μl drop of a 10⁸ cfu/ml bacterial suspension. After eight weeks’ incubation in a greenhouse at 22 ±1°C, bleeding cankers developed at the inoculation sites of all treated trees, while control trees remained symptomless (Fig. 3). The average lesion lengths produced by isolates M8597 and M8603 were 38.8 mm (range 18-67 mm) and 31.4 mm (range 21-52 mm), respectively. Koch’s postulates were completed by successfully re-isolating the bacteria from all the necrotic lesions.

This is the first detection of P. syringae pv. aesculi on horse chestnut trees in Switzerland. The bleeding canker symptoms are moderate and being monitored to follow their progression. Since the outbreak of the disease in Northern Europe (UK and Netherlands) in the early 2000’s (Webber et al., 2008), the pathogen has been found in other countries causing economic losses (EPPO, 2009). Isolates M8597 and M8603 have been deposited in the Culture Collection of Switzerland (CCOS) as CCOS950 and CCOS951, respectively.