Genomic diversity of a Microviridae phage CLasMV1 in “Candidatus Liberibacter asiaticus”

Authors: WANG, CHENG - South China Agricultural Univerisity; FANG, FANG - South China Agricultural Univerisity; ZHENG, MINGXIN - South China Agricultural Univerisity; ZHENG, YONGQIN - South China Agricultural Univerisity; GAO, CHENYING - South China Agricultural Univerisity; Wallis, Christopher; LI, ZIYI - South China Agricultural Univerisity; Chen, Jianchi; DENG, XIAOLING - South China Agricultural Univerisity; ZHENG, ZHENG - South China Agricultural Univerisity

Submitted to: Phytopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/30/2025
Publication Date: 6/4/2025
Citation: Wang, C., Fang, F., Zheng, M., Zheng, Y., Gao, C., Wallis, C.M., Li, Z., Chen, J., Deng, X., Zheng, Z. 2025. Genomic diversity of a Microviridae phage CLasMV1 in “Candidatus Liberibacter asiaticus”. Phytopathology. https://doi.org/10.1094/PHYTO-04-25-0133-SC.
DOI: https://doi.org/10.1094/PHYTO-04-25-0133-SC

Interpretive Summary: Citrus Huanglongbing (HLB) is a destructive disease affecting citrus production around the world. The disease is caused by the infection of a bacterial pathogen. Study on HLB pathogen is difficult because it cannot be cultured in artificial media. Using recent genomic technology, a virus called CLasMV1 that infects the HLB pathogen was discovered. This study was to characterize the bacterial virus using 1,042 samples collected from China where HLB was first described more than a hundred years ago. Forty-three viral genome sequences were analyzed and used to build a classification system to meet current research need. The new information obtained from this study expands our knowledge on HLB pathogen that will help the disease management.

Technical Abstract: A Microviridae phage, CLasMV1was recently identified in “Candidatus Liberibacter asiaticus” (CLas), a nonculturable bacterium associated with citrus Huanglongbing (HLB). The representative CLasMV1 genome (CP045566, Strain GDHZ11D) is circular with 8,869 bp including eight open reading frames (ORFs). Multiple CLasMV1 sequences have now been identified in China, yet knowledge about the phage biology, such as gene functions and population diversity are limited. In this study, 1,520 CLas samples were collected from ten citrus-growing provinces in China. PCR experiments identified 1,042 samples (69%) harboring CLasMV1. Complete genome sequences of 43 CLasMV1 strains were obtained through Illumina HiSeq sequencing and reference-mapping to CP045566. The phage genome size ranged from 8,696 bp to 8,881 bp. The viral genome variation is associated with single-nucleotide polymorphisms (SNPs) and sequence insertion/deletions (In/Dels). Two hypervariable regions (HVRs) were detected, HVR-1 spanning from position 1,547 to 1,729 (182 bp), and HVR-2 spanning from position 4,603 to 5,014 (411 bp). Sequence analysis using 43 CLasMV1 genomes from China and two homologs from Florida CLas strains (CP040636, MZ382797) revealed five genomic clusters. A primer set flanking HVR-1 region was designed and used to test 1,042 CLasMV1 samples. A 174-bp insertional/deletional sequence was detected, suggesting the presence of mobile element in CLasMV1. PacBio long-read sequencing experiment detected concatemer of CLasMV1 genome, suggesting the use of rolling-cycle mechanism in the viral replication. New information obtained from this study expanded our knowledge of CLasMV1 phage and will facilitate future research on CLas phages for better understanding of HLB biology and management.