First Report of Xanthomonas albilineans Causing Leaf Scald on Two Chewing Cane Clones in Zhejiang Province, China

Abstract

In recent years, outbreaks of leaf scald have been reported in two chewing cane clones “Guangdong Huangpi” and “Taoshang Guozhe” in Zhejiang province, China. From May to July 2019, we collected 11 and 13 leaf or stalk samples from symptomatic “Guangdong Huangpi” from four farms in Wenling and “Taoshang Guozhe” clones from three farms in Ruian, Zhejiang province, respectively. Leaves in young plants exhibited white pencil-line streaks (Supplement Fig. 1A & 1D) as well as partial or complete chlorosis of the leaf blade (Supplement Fig. 1B & 1E). Internal symptoms included an orange-red discoloration of the vascular bundles at the basal nodes of the stalk and discoloration extension into the internodes (Supplement Fig. 1C & 1F). Leaf and stalk tissues were used for bacterial isolation and purification on XAS medium, which is selective for Xanthomonas albilineans (Davis et al. 1994), using the streak plate method to obtain 24 isolates (Lin et al. 2018). Circular, convex, smooth, shiny and yellow colonies were isolated from all the samples. The pathogenicity of two isolates, XaCN30 from “Guangdong Huangpi” and XaCN43 from “Taoshang Guozhe”, was confirmed with Koch’s postulates according to the protocol reported by Lin et al. (2018). The incidences of diseased plants (56% and 63%) were observed in individual host clones at 28 d post-inoculation with isolates XaCN30 and XaCN43, respectively. Furthermore, all isolates were confirmed as X. albilineans via molecular methods. PCR amplification was conducted for all 24 isolates using the primer pairs XgyrB1F/XgyrB1R2 (Ntambo et al. 2019) and XAF1/XAR1 (Wang et al. 1999), which targeting the gyrB (encoding the b subunit of the DNA gyrase) and abc (encoding an ABC transporter) genes, and generating 904 bp and 608 bp amplicons, respectively. The PCR fragments were cloned into the pMD19-T vector (TaKaRa, Dalian, China). For each isolate, three single colonies of transformed Escherichia coli DH5α carrying targeted fragment were sequenced. These sequences were deposited into the GenBank with accession no. MT776053-MT776059 and MT776061-MT776077 for gyrB gene and MT776098-MT776104 and MT776106-MT776122 for abc gene. Based on the two concatenated DNA sequences of our 24 isolates, compared with 27 previously reported X. albilineans isolates obtained from the GenBank database, pairwise sequence identity analysis revealed that all 24 isolates from Zhejiang province had 99.4-100% identity with each other, 99.6-100% identity with 14 published domestic isolates, and 98.3-100% identity with 13 foreign isolates. Furthermore, phylogenetic analysis with MEGA 7.0 (Kumar et al. 2016) showed that the isolates from Zhejiang province clustered into two distinct groups (Supplement Fig. 2). One group consisted of 25 Chinese isolates (including all 11 isolates from Wenling) along with four isolates from the French West Indies (GPE PC73, GPE PC17, GPE PC86, and MTQ032), and one isolate from the USA (XaFL07-1), which were assigned to pulsed-field gel electrophoresis (PFGE) group B (Davis et al. 1997; Pieretti et al. 2012). A putative group was also proposed, which included all 13 isolates from Ruian, indicating that isolates from Ruian are distinct from the isolates isolated from other Chinese sugarcane-planting areas, including Wenling. We conclude that leaf scald disease in local clones of chewing cane are caused by X. albilineans in Zhejiang province in China, which will be helpful for leaf scald management in chewing cane, a cash crop.

Genetic Divergence and Population Structure of Xanthomonas albilineans Strains Infecting Saccharum spp. Hybrid and Saccharum officinarum

“…hybrid), grown within China. However, some Xa strains were collected from a local traditional chewing cane cultivar Taoshangguozhe ( S. officinarum ), collected in the city of Ruian (Zhejiang province), and were proposed to form a new phylogenetic group, Xa-RA, based on MLSA with two ( ABC and gyr B) [ 22 ] or four ( rpo D, gln A, gyr B, and atp D) genes [ 7 ].…”

“…The strains hosted in the cultivar Taoshangguozhe of S. officinarum formed a separate group, group II, which was distinct from other Chinese strains, whereas the Xa strains hosted in two chewing cane cultivars (Guangdonghuangpi and Badila) of S. officinarum along with Xa strains hosted in modern commercial cultivars clustered into the same group (Group I). Notably, the famous chewing cane cultivar Badila has been cultivated for more than half a century in southern China, while the cultivar Guangdonghuangpi was recently introduced into Zhejiang from Guangdong province [ 22 ]. Almost all Xa strains from China, including those grown in Guangdong province, clustered in the PFGE-B group [ 21 , 30 ].…”

“…Pulsed-field gel electrophoresis (PFGE) and multilocus sequence analysis (MLSA) revealed that 218 isolates from 31 geographical locations were clustered into 10 genetic groups (PFGE-A to PFGE-J) [ 18 , 19 , 20 ]. Our recent studies revealed that the majority of Xa strains in China belonged to PFGE-B [ 21 ], while some strains from a novel group were proposed based on MLSA and simple sequence repeats (SSRs) [ 7 , 22 ]. In addition to the MLSA and SSR methods, other fingerprinting tools have also been used to investigate Xa genetic divergence, including restriction fragment length polymorphism (RFLP) [ 18 ], random amplified fragment polymorphism (RAPD) [ 23 ], rep-PCR [ 24 ], and amplified fragment length polymorphism (AFLP) [ 25 ].…”

“…Our previous studies examined the genetic diversity of Xa in China based on MLSA and SSR techniques [ 7 , 21 , 22 ], although the population divergence and evolution of this pathogen remain largely unexplored. The aims of this study were to: (a) use rep-PCR and MLSA to investigate the genetic diversity of 39 Xa strains from China, (b) estimate the population structure of these strains, and (c) analyze the evolutionary driving force that forms Xa populations from a modern commercial cultivar ( Saccharum spp.…”

Sugarcane responses to two strains of Xanthomonas albilineans differing in pathogenicity through a differential modulation of salicylic acid and reactive oxygen species

“…Up to now, only X. albilineans strains clustering with foreign strains of PFGE group B were reported in China ( Zhang et al., 2017 ; Ntambo et al., 2019b ). Nevertheless, recent observations suggested that other phylogenetic groups of X. albilineans occur in China ( Duan et al., 2021 ).…”