Characterization of Erwinia amylovora causing Twig Blight of Loquat Tree in Loquat Growing Areas of Punjab and KPK, Pakistan

Abstract

Twig blight of loquat (Eriobotrya japonica Lindl.), caused by the gram-negative enterobacterium Erwinia amylovora, is a major constraint to loquat production worldwide. Recent surveys in loquat-growing areas of Punjab and KPK, Pakistan—specifically in Chakwal (Kalar Kahar, Choa Saidan Shah), Rawalpindi (Taxila, Wah), Murree (Tret and Chattar), and Haripur (Haripur, Khanpur, and Sara i Saleh)—revealed that this disease is highly prevalent. Twig blight compromises the health and vigor of loquat trees, often leading to their death, which discourages farmers from cultivating loquat commercially. Consequently, the area under loquat cultivation and its production have both been declining. Despite the severity of the issue, no systematic studies had been conducted to address this problem until now. A survey was carried out over two consecutive years (2016 and 2017) in 10 loquat-growing locations in Punjab and KPK (three orchards per location). The incidence of twig blight was higher in 2017 compared to 2016. The highest incidence was recorded in Taxila, with 69.74% in 2016 and 78.39% in 2017, while the lowest was in Kalar Kahar (20.86%). The incidence in Kalar Kahar showed a slight increase of less than 1% from the previous year, likely due to effective control measures implemented to limit the spread of the disease. The pathogen E. amylovora was isolated from diseased samples. Out of 102 bacterial isolates, 92 strains were gram-negative, and 72 tested positive in various biochemical assays (Levan, lipase, KOVAC oxidase, glucose fermentation, catalase oxidase, growth at 39°C, production of fluorescent pigment, and pectolytic activity tests). Three highly virulent isolates were identified through pathogenicity tests and confirmed by biochemical and molecular indicators. Molecular tagging of two chromosomal genes (amsB [MN902191] and Pst-glms [MN915084]) and one plasmid gene (pAE29 [MN902192]) was performed, confirming the identity of the isolates as E. amylovora. This study represents the first report of E. amylovora infecting loquat in Pakistan.