Prophage Diversity of ‘CandidatusLiberibacter asiaticus’ Strains in California

Abstract

Huanglongbing (HLB) is a highly destructive citrus disease and is associated with a nonculturable bacterium, ‘Candidatus Liberibacter asiaticus’. ‘Ca. L. asiaticus’ in the United States was first found in Florida in 2005 and is now endemic there. In California, ‘Ca. L. asiaticus’ was first detected in Hacienda Heights in Los Angeles County in 2012 and has now been detected in multiple urban locations in southern California. Knowledge of ‘Ca. L. asiaticus’ strain diversity in California is important for HLB management. In this study, genomic diversity among 10 ‘Ca. L. asiaticus’ strains from six California locations were analyzed using a next-generation sequencing (NGS) (Illumina MiSeq and HiSeq) approach. Draft genome sequences of ‘Ca. L. asiaticus’ strains were assembled. Sequences of the 16S ribosomal RNA gene and nrdB confirmed ‘Ca. L. asiaticus’ identity. Prophages were detected in all ‘Ca. L. asiaticus’ strains. The California ‘Ca. L. asiaticus’ strains formed four prophage typing groups (PTGs): PTG1, with type 1 prophage only (strains from Anaheim, San Gabriel, and Riverside); PTG2, with type 2 prophage only (strains from Hacienda Heights); PTG1-3, with both type 1 and 3 prophages (a strain from Cerritos); and PTG1-2, with both type 1 and type 2 prophages (a strain from La Habra). Analyses of the terL sequence showed that all California ‘Ca. L. asiaticus’ strains were not introduced from Florida but likely from locations in Asia. Miniature inverted-repeat transposable elements were found in all ‘Ca. L. asiaticus’ strains, yet, a jumping-out event was detected in the ‘Ca. L. asiaticus’ strain from Cerritos. Altogether, this study demonstrated that the NGS approach focusing on prophage variation was sensitive and effective in revealing diversity of ‘Ca. L. asiaticus’ strains in California.

Occurrence of prophage and historical perspectives associated with the dissemination of huanglongbing in mainland China

“…DNA mosaicism and hypersequence variation were identified and successfully used to differentiate CLas strains from Yunnan and Guangdong following PCR of several prophage genes and the documentation of dominant electrophoretic profiles (E‐types; Wang et al , ). A Type 3 prophage was discovered recently (Zheng et al , ) and employed for diversity analysis of CLas strains in California and southern China (Dai et al , ; Li et al , ).…”

Functional and comparative genomic analysis of integrated prophage-like sequences inCandidatus Liberibacter asiaticus

“…[15,11,10], whether of Type 1, 2, or 3. These phage types can be used to compare evolutionary dynamics [12,9], and to predict the origin of strains [17,14,9]. Nonetheless, prior work has leveraged read-mapping based approaches to identify new prophage-like sequences, albeit those that resemble Type 1 or 2 phage.…”

“…Likewise, the last part of the AHCA1-a sequence and the following region found after a genome assembly gap reconstruct a Type 3 prophage with the sequence organization predicted for P-JXGC-3 [13] (Figure 1b). Only a Type 1 prophage has been reported in strain AHCA1 [9], but the sequence organization of the integrated prophage ( Figure 1) and the presence of a hypothetical protein (YP 007011137.1) only found in prophages of Type 2 and 3 (see Supl. Table 3), suggest the CLas strain AHCA1 harbours a Type 3 prophage sequence.…”

“…The CLas genome includes genes involved in cell motility and active transport [5,6], despite the presence of these genes, only passive movement has been observed in the phloem sap [6]. Furthermore, prophage sequences have also been identified in multiple CLas strains [7,8,9,10,11,12,13,14,15]. The interest in Clas prophage stems from observations that a prophage-encoded peroxidase is an effector that suppresses plant defenses [3].…”

“…Type 1, 2 or 3. Current methods to identify prophage in CLas genomes rely on read mapping against known prophages [12,9]. As a consequence, failure of read-mapping has been interpreted to mean that a genome is prophage free (e.g., as in the Ishi-1 strain [16]).…”