Tomato brown rugose fruit virus (ToBRFV), first identified in 2014 in the Middle East, has caused serious disease outbreaks on greenhouse tomatoes in many countries in Asia, Europe and North America. With its seed-borne, easy mechanical transmission and resistance breaking to the popular Tm-22 gene, ToBRFV could potentially cause serious economic losses to tomato industries worldwide if not managed timely. In the U.S., the first ToBRFV outbreak on tomato plants was identified in 2018 in a greenhouse facility in California. The objective of this study is to characterize the molecular and biological properties of this emerging ToBRFV isolate and to achieve a sensitive, species-specific detection. In the current study, high throughput sequencing is used to obtain a complete genome sequence of the US isolate of ToBRFV. The strong nucleotide sequence identity (99%) to other known ToBRFV isolates around the world suggests a high possibility of sharing the same or similar origin. Comparative evaluation of potential host ranges among tomato-infecting tobamoviruses (ToBRFV, tomato mosaic virus and tomato mottle mosaic virus) showed many similarities, but some unique plant species were also identified for each respective virus. Due to serological cross reactivity among tomato-infecting tobamoviruses, a species-specific real-time RT-PCR was developed, which was shown to provide sensitive and specific detection of ToBRFV.