Identification of a Novel 1033‐Nucleotide Deletion Polymorphism in the Prophage Region of ‘Candidatus Liberibacter asiaticus’: Potential Applications for Bacterial Epidemiology

Abstract

The prophage/phage region in the genome of ‘Candidatus Liberibacter asiaticus’, an alpha‐proteobacterium associated with citrus Huanglongbing, included many valuable loci for genetic diversity studies. Previously, a mosaic genomic region (CLIBASIA_05640 to CLIBASIA_05650) was characterized, and this revealed inter‐ and intracontinental variations of ‘Ca. L. asiaticus’. In this study, 267 ‘Ca. L. asiaticus’ isolates collected from eight provinces in China were analysed with a primer set flanking the same mosaic region plus downstream sequence. While most amplicon sizes ranged from 1400 to 2000 bp, an amplicon of 550 bp (S550) was found in 14 samples collected from south‐western China. Sequence analyses showed that S550 was the result of a 1033 bp deletion which included the previously known mosaic region. The genetic nature of the deletion event remains unknown. The regional restriction of S550 suggests that the ‘Ca. L. asiaticus’ population from south‐western China is different from those in eastern China. The small and easy‐to‐detect S550 amplicon could serve as a molecular marker for ‘Ca. L. asiaticus’ epidemiology.

Genetic Diversity of “ Candidatus Liberibacter asiaticus” Based on Four Hypervariable Genomic Regions in China

“…The genetic diversity of uncultured C Las bacteria is primarily accessed by amplifying the conserved genes, such as the 16S rRNA gene, 16S/23S rRNA intergenic spacer regions ( 11 ), the outer membrane protein gene ( omp ) ( 12 ), the deoxy-ribonucleotide reductase gene ( nrdB ) ( 13 ), the β-operon gene loci ( 14 ), the tufB–secE–nusG–rplKAJL–rpoB gene cluster ( 15 ), tandem-repeat ( 16 to 18 ), or hypervariable prophage regions ( 19 to 21 ). Genetic variations within these analyzed nonprophage regions were low and could hardly distinguish closely related C Las.…”

“…Genetic variations within these analyzed nonprophage regions were low and could hardly distinguish closely related C Las. C Las does have several hypervariable prophage regions ( 19 to 22 ). However, the prophage varies from 0 to 2 copies in different C Las isolates and could only distinguish partial C Las isolates ( 23 to 25 ).…”

Graft-transmissible citrus diseases in P. R. China-research developments

“…In the P.R. China, ten graft-transmissible pathogens have been identified infecting citrus, including Candidatus Liberibacter asiaticus, Citrus tristeza virus (CTV), Citrus tatter-leaf virus (CTLV), Citrus exocortis viroid (CEVd), Citrus yellow vein clearing virus (CYVCV), Satsuma dwarf virus (SDV), Citrus vein enation virus (CVED), Citrus psorosis virus (CPsV), Citrus cachexia viroid (CCaVd) and Citrus chlorotic dwarf virus (CCDV) (Fu et al 2015;Huang et al 2015;Lin et al 2013;Lin et al 2015;Shang et al 2016;Song, Kurth, et al 2015;Wang, Su, et al 2015;Wang et al 2013;Wu et al 2014;Yi and Zhou 2014;Zheng et al 2014;Zheng and Zhou 2013;Zhou et al 2017;Zhou et al 2013). Of these pathogens, the first five cause damage in field citrus trees, whereas the latter five were occasionally detected from the imported citrus materials or field trees.…”

“…Some progress has been made: 1) putative prophage particles were observed in sweet orange (Fu et al 2015); 2) the whole genomes of three Chinese isolates of the HLB bacterium were sequenced; bio-information is accumulating quickly, focusing on the prophage genomic region with high genetic variation and recombination events; two HLB origin centres were proposed in the P.R. China (Lin et al 2013;Lin et al 2015;Wang et al 2013;Wang, Su, et al 2015;Zheng et al 2014;Zheng and Zhou 2013); 3) transgenic citrus lines against HLB are in field trials for evaluation; 4) a few effective combinations of pesticides have been selected to control citrus psyllids (Diaphorina citri); 5) natural thermotherapy of the citrus tree canopy by covering with PVC mulch showed somewhat efficacy to reduce the disease symptoms.…”