First Report of Shoot Blight on Fragrant Pear Caused by Pantoea ananatis in China

Abstract

Fragrant pears are important economic plants in Xinjiang region, a major agricultural production area in China. In March 2017, a strange shoot blight disease on fragrant pear was initially observed in Kolar, Xinjiang. The incidence of the disease was up to 50% in certain orchards, and the total incidence area was approximately 26,000 ha of commercial fields. The disease developed from blossoms to shoots and larger branches. More disease was observed on old and wounded trees. Symptoms included wilted shoot and leaves. When infections spread to larger tree limbs, the bark of branches became darkened and water-soaked. After bark shaving, dark xylem tissue was visible. Shoot samples were collected from diseased plants (Pyrus sinkiangensis Yu) with characteristic symptoms. The shoot samples were surface sterilized with 75% ethanol and 1% sodium hypochlorite, rinsed three times in sterilized water, cut into small pieces, and then soaked in 10 ml of sterile phosphate buffered saline (PBS) for 30 min. The suspension was streaked onto a nutrient agar plate and incubated at 30°C. After 48 h, dominating light yellow, round flat colonies with smooth surfaces were picked and restreaked for purification. The bacteria were identified by polymerase chain reaction amplified 16S rRNA gene and further by multilocus sequence analysis of partial nucleotide sequences of the genes rpoB and infB using rpoB CM7-F/CM31b-R (1,000 bp) and infB 01-F/02-R (1,030 bp) primer pairs, respectively (Brady et al. 2008). BLAST analysis showed that the 16S rRNA (NCBI accession no. MK128507), rpoB (MK135821), and infB (MK135822) gene sequences shared 100, 100, and 99% similarity to that of Pantoea ananatis strains BSP9 (KF360069.1), 15320 (MH015163.1), and BRT98 (MH015108.1), respectively. In April 2018, 3-year-old fragrant pear trees were inoculated with the P. ananatis in a separated garden. The branches of trees were injured with cuts of 2 to 3 cm in the bark. Autoclaved cotton balls were soaked with bacterial suspensions of 10⁸ CFU/ml and attached to the wounds with Parafilm. The bacterial suspensions were inoculated on three trees, and an additional two trees were treated with PBS instead as a negative control. Three shoots were inoculated per tree. The trees were then grown under natural conditions. After 12 weeks, symptoms resembling bacterial shoot blight symptoms had developed on all inoculated shoots. Control trees did not develop symptoms. Reisolations were performed from symptomatic tissues, and isolates were identified as P. ananatis using the techniques for molecular identification as mentioned above. P. ananatis was initially discovered on pineapple causing fruitlet rot in the Philippines in 1928 (Serrano 1928), and it is regarded as an emerging pathogen based on the increasing number of reports of diseases occurring on previously unrecorded hosts in different parts of the world. Coutinho et al. (2002) reported similar symptoms including shoot wilt and die-back after the infection of Eucalyptus trees with P. ananatis. In 2017, isolation and identification of brown rot bacteria P. ananatis was reported on imported succulent plants in China (Li et al. 2017). To our knowledge, this is the first report of P. ananatis causing shoot blight on fragrant pear in China. This study expanded the host range and geographic location of P. ananatis as a plant pathogen and suggests that commercial fragrant pear producers should take early control actions to prevent spread of and reduce losses caused by the disease.