Cadmium is a highly toxic heavy metal for biological systems. Cupriavidus metallidurans CH34 is a model strain for heavy metal resistance and bioremediation. The aim of this study was to determine the role of the c-di-GMP pathway in the C. metallidurans CH34 response to cadmium in both planktonic and biofilm cells. Increasing cadmium concentrations correlates with an inhibition of biofilm formation and EPS production in C. metallidurans cells. Planktonic and biofilm cells showed similar tolerance to cadmium. During exposure to cadmium an acute decrease of c-di-GMP levels in planktonic and biofilm cells was observed. Transcription analysis by RT-qPCR showed that cadmium induced in planktonic cells and strongly induced in biofilm cells the expression of the urf2 gene and the mercuric reductase encoding merA gene, which belong to the Tn501/Tn21 mer operon. After exposure to cadmium the cadA gene involved in cadmium resistance was equally upregulated in both lifestyles. Bioinformatic analysis and null mutant complementation assays indicated that the protein encoded by the urf2 gene is a functional phosphodiesterase involved in the c-di-GMP metabolism. We propose to rename the urf2 gene as mrp gene for metal regulated phosphodiesterase. An increase of the second messenger c-di-GMP content by the heterologous expression of the constitutively active diguanylate cyclase PleD* correlated with an increase in biofilm formation and cadmium susceptibility. These results indicate that the response to cadmium in C. metallidurans CH34 involves a decrease in c-di-GMP content that inhibits the biofilm lifestyle.

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.