First Report of Hoja Blanca Viral Disease in Rice in Texas, U.S.A.

Hoja Blanca is a major disease affecting rice (Oryza sativa L.) production in Central and South America and the Caribbean, with yield loss reaching up to 75% (Morales and Jennings 2010). The disease is caused by the Rice hoja blanca virus (RHBV), a member of the genus Tenuivirus, and is transmitted by the planthopper Tagosodes orizicolus Muir. In September 2024, symptoms resembling those of Hoja Blanca were observed in rice ratoon crops across 1,133 hectares in Waller and Wharton Counties, Texas, following outbreaks of planthoppers. The infected plants exhibited chlorotic stripes or mottled discoloration on the upper leaves, stunted growth, flower sterility, and poor grain filling (Fig. S1). Affected rice cultivars included CLL18, CLL153, CLJ01, Dixiebelle, Presidio, PVL03, and RT7302 (hybrid). Symptomatic plant samples were collected, including two each from Waller County (24-TX-Br1 and 24-TX-Br2) and Wharton County (24-TX-Wh1 and 24-TX-Wh2). The presence of the virus was tested using the commercial RHBV ELISA kit following the manufacturer’s protocol (Creative Diagnostics, NY, USA). The samples were then subjected to virus confirmation via sequencing. Total RNA was extracted from 50 mg of leaf tissue using the Quick-RNA miniprep plus kit (Zymoresearch, CA, USA) and cDNA was synthesized from 2 µg of RNA using the PrimeScript 1st strand cDNA synthesis kit (Takara Bio, CA, USA). This cDNA served as template for the PCR amplification using the following primers: RHBV-R3-F: 5′-CCTTTCCAGTTTCATCTACCATC-3′ and RHBV-R3-R: 5′-AACAATGACGATGCCAGTTGCTGA-3′, and RHBV-R4-F: 5′-CTCAGGCCTCAATCATTACC-3′ and RHBV-R4-R: 5′-ACTTTCAGGATAATAGACAG-3′ (Bolaños et al. 2017). Amplicons of the expected sizes of 980 bp and 466 bp, respectively, were visualized, purified (Zymo Research, CA, USA), and subjected to sanger sequencing (Eton biosciences, NC, USA). Sequence analysis of the RNA3 region from all four samples – 24-TX-Br1 (GenBank: PQ560683), 24-TX-Br2 (PQ560684), 24-TX-Wh1 (PQ560681), and 24-TX-Wh2 (PQ560682) – revealed >90% similarity to other RHBV isolates previously reported from Colombia (MG566076, AF004658), Costa Rica (L07940), and Peru (KY369921). Similarly, RNA4 region for all four samples – 24-TX-Br1 (PQ560685), 24-TX-Br2 (PQ560686), 24-TX-Wh1 (PQ560687), and 24-TX-Wh2 (PQ560688) – shared >95% similarity to other RHBV isolates previously reported from Colombia (MG566077, L14952), Costa Rica (AF004657), and Peru (KY369922). Phylogenetic analysis confirmed that all four virus isolates clustered with other reported RHBV isolates, forming a distinct clade that was separate from other Tenuivirus species. To fulfill Koch’s postulates, six one month old plants (cv. Presidio) were individually infested with 10 viruliferous plant hoppers for three days in a cage, after which the insects were removed. After 30 days, RHBV symptoms appeared on infested plants but not on the control plants. The presence of RHBV in symptomatic plants was confirmed using ELISA and PCR. In the U. S., Hoja Blanca was first reported in Florida in 1957 (Atkins and Adair, 1957), in Mississippi in 1958 and Louisiana in 1959 (Atkins 1966) but has since disappeared. To our knowledge, this is the first report of Hoja Blanca in Texas, marking the first virus disease affecting rice in the U.S. This underscores the urgent need to establish surveillance systems to monitor the potential spread of this invasive virus and its insect vectors across Texas and other rice-producing regions.