This study examined the efficiency of a combined diagnostic approach in which bioamplification of Arizona 861 S1 citron was integrated with RT‒PCR to detect Citrus tristeza virus (CTV) and Hop Stunt Viroid (HSVd) in Citrus plants. The capacity of the combined method to diagnose mixed infections of CTV and HSVd, along with early single infections of both pathogens, was assessed by evaluating the incubation periods and differential detection capabilities between the combined and direct methods. The bioamplification method exhibited high sensitivity (100%) and specificity (100%) for CTV, enabling early detection even in patients with low-titer infections compared to direct RT‒PCR. However, for HSVd, the combined method achieved 50% sensitivity in early infections, highlighting the need for additional replicates to enhance diagnostic reliability. The relative concentrations of pathogens in mixed and simple infections were evaluated; the analysis revealed no significant differences, suggesting minimal interference between CTV and HSVd. This approach allows early detection of CTV but requires further refinement for HSVd diagnosis. These findings underscore the complementary roles of the cost-effectiveness of direct RT‒PCR and the sensitivity of the combined method in implementing effective disease control strategies in citrus crops.

Grapevine trunk diseases (GTDs) attack the vine's wood, devastating vineyards worldwide. Chile is the world's fourth largest wine exporter, and Cabernet Sauvignon leads the red wine varieties. Botryosphaeria dieback is an important GTDs and Diplodia seriata is one of the main pathogenic species. Biocontrol studies of these pathogens are commonly carried out at different incubation times but at a single temperature. This study aimed to evaluate the biocontrol effect of native Chilean bacteria against D. seriata isolates at different temperatures. We analyzed the biocontrol effects of Pseudomonas sp. GcR15a, Pseudomonas sp. AMCR2b and Rhodococcus sp. PU4, with three D. seriata isolates (PUCV 2120, PUCV 2142 and PUCV 2183) at 8, 22 and 35°C. Agar plug diffusion and double plate methods were used to evaluate the effect of diffusible and volatile organic compounds (DOC and VOC), and an in vivo test was performed with Cabernet Sauvignon cuttings. Pseudomonas strains showed the most significant inhibition in the DOC assay, and a demelanizing activity was observed. Rhodococcus sp. PU4 showed VOC biocontrol potential at high temperatures (35°C). In vivo, a biocontrol effect was observed against two D. seriata isolates, while the PUCV 2142 isolate proved to be more resistant to being controlled. The biocontrol ability at low temperatures is promising for effective control in the field, where infections occur primarily in winter.

In vineyards, Botryosphaeria dieback (BD) occurs in young and old plants. In the field, the prevalence and severity of the disease increase proportionally with the age of the vineyards. Among the pathogens that cause (BD), Diplodia seriata is the most prevalent species in Chile and other countries with a Mediterranean climate. To date, no information is available on the sus-ceptibility of adult wood to infection by this pathogen since most of the pathogenicity tests have been carried out on 1- or 2-year-old shoots or detached canes. Therefore, a pathogenicity test was carried out on plants under field conditions, with inoculations in plants, on one-year-old shoots, and 2- and 10-year-old wood in grapevine cv. Cabernet Sauvignon. A pathogenicity test was carried out with two isolates of D. seriata selected in an assay in detached canes that were 2-year-old. The results on plants showed that D. seriata was significantly more aggressive on 10-year-old than on one- or two-year-old tissue. These results were compared with the lesions ob-tained from two-year-old canes. The results of this work are consistent with the damage ob-served in the field, highlight the importance of identifying each pathogenic species that causes (BD) and contribute to the knowledge of the epidemiology of this disease in Mediterranean cli-mates.