The Phylogeographic History of Pepper Mild Mottle Virus

Pepper mild mottle virus (PMMoV) belongs to the species Tobamovirus capsicai in the genus Tobamovirus (family Virgaviridae), has emerged as an important pathogen, significantly impacting pepper yield and quality. The objectives of this study are to investigate its phylogeographic history and evolutionary dynamics, and to lay the foundation for accurate monitoring and scientific prevention and control

A specific primer pair flanking the PMMoV coat protein (CP) gene was designed. The CP gene sequences of 28 randomly selected PMMoV isolates from Fujian Province were subsequently amplified and cloned. In addition to the newly obtained sequences, all publicly available CP sequences from GenBank with known collection timestamps and geographical origins were assembled into a final dataset of 255 sequences. After confirming a sufficient temporal signal via a date-randomized test (DRT), a structured coalescent-based Bayesian phylodynamic framework (MultiTypeTree) was employed to reconstruct the evolutionary history and spatial dissemination of the virus.

A target fragment with the expected size was obtained from all 28 PMMoV-positive samples selected in this study. Their CP sequences share＞98% nucleotide identity with known PMMoV isolates. The DRT showed no overlap in the 95% confidence intervals of substitution rates between the real and randomized datasets, confirming a reliable temporal signal for Bayesian molecular dating. Phylogenetic analysis estimated a mean substitution rate of 9.24×10^-4 substitutions/site/year (95% CI: 6.20×10^-4-1.01×10^-3 substitutions/site/year) for the PMMoV CP, which is comparable to rates observed in animal RNA viruses, indicating its rapid evolutionary dynamics. The time to the most recent common ancestor (tMRCA) was dated to 1941 (95% CI: 1921-1957). The root of the maximum clade credibility (MCC) tree was placed in Europe, identifying this region as the most probable origin of contemporary global PMMoV isolates. Temporal migration analysis revealed multiple dispersal routes from Europe to other regions, underscoring its role as a central hub in the virus’s global dissemination. In addition to cross-regional transmission, frequent local spread of PMMoV was observed within regions. Bayesian skyline plot (BSP) analysis further revealed distinct historical population dynamics: the South American population underwent a significant expansion before stabilizing, whereas those in most other regions remained relatively stable over the long term.

Europe served as the most likely source and a critical dissemination hub for the global spread of PMMoV. This study provides a foundation for understanding the molecular epidemiology of PMMoV and informs the development of future disease management strategies.