A total of 116 grape samples (50 summer grape disease samples and 66 winter grape disease samples) were collected from 21 orchards across five regions in three major grape-producing counties. In these regions, Kyoho grapes are the predominant variety, occupying the largest planting area. However, a few other grape varieties were found in limited cases on some farms. Due to the presence of browning compounds, such as polyphenols in grape leaves, the extraction of viral RNA from grapes is often interfered with. Therefore, developing a reliable nucleic acid extraction method for grape viruses is essential. Two commonly used nucleic acid extraction methods that the semi-automated extraction instrument (TANBead) and the RNeasy Plant Mini Kit (Qiagen) were tested. Nucleic acids were extracted from six samples using both methods, and the RNA quality was evaluated based on the 260/280 and 260/230 ratios, as well as the amplification of endogenous grape genes. Results indicated that the semi-automated extraction instrument (TANBead) yielded superior nucleic acid quality. The purified RNA samples were subsequently subjected to small RNA next generation sequencing (NGS). In addition, the collected materials were tested using five serological diagnostic methods previously established in the laboratory to detect Grapevine virus A (GVA), Grapevine fanleaf virus (GFLV), Grapevine leafroll-associated virus 1 and 3 (GLRaV-1 and GLRaV-3), and Grapevine fleck virus (GFkV) via ELISA assays. Preliminary results on virus infections revealed that GFkV was the most frequently detected virus, either as a single infection or in mixed infections. However, GFLV was not detected in any sample. In Fengqiu, all 21 samples tested positive for viruses, including 7 samples of single-virus infections, 13 samples of mixed infections, and 1 sample of triple-virus infection. It is hypothesized that the cultivation and management practices in this region may contribute to the high frequency of viral infections. According to Yang et al. (2000), a two-year survey on the reinfection of tissue culture seedlings by viruses in the field revealed the presence of GLRV-1 in Taiwan, along with the detection of GFLV and GVA. However, this differs slightly from the survey results of the current year (2024), where no GFLV was detected, but GLRaV-3 was identified in the collected samples. This indicates a shift in the composition of grapevine viruses present in the field.

▲Fig. 1 Collection of disease materials from different grape varieties in major domestic production areas.		▲Fig. 2 The nucleic acid of grapes is subjected to PCR reaction using the endogenous gene RRM1 primer pair (reference genes) of grape plants to confirm the quality of the nucleic acid samples.
▲Fig. 3 A total of 116 samples were tested using ELISA serological methods for five grape viruses (GVA, GFLV, GLRaV-1, GLRaV-3, GFkV).		▲Fig. 4 The infection status of grapes in various regions corresponds to specific viruses.