Botryosphaeria disease in New Zealand blueberry gardens: identification of pathogens, inoculum sources and factors affecting disease development

Abstract

Botryosphaeriaceae species cause dieback and death of blueberry plants but the species involved and factors affecting infection and spread are not well understood. Sampling at blueberry farms found these pathogens in five of seven farms sampled; overall incidence was 41.4%, with N. australe (79.0%), N. luteum (8.0%), N. ribis (8.0%) and N. parvum (5.0%). Sampling of nursery plants found infections in all four nurseries with 45% incidence in mainly asymptomatic plants, which were infected with N. australe (66.0%), N. parvum (31.5%) and N. ribis (2.5%). Asymptomatic propagation cuttings from one nursery were found to have external contamination of Botryosphaeriaceae DNA (90.0%) and internal infection (65.0%) by the main four species found in the blueberry farms and nurseries. Propagation media received from four nurseries contained Botryosphaeriaceae DNA (44.0% incidence), with species identified using SSCP as N. australe, N. luteum, N. parvum/ N. ribis and D. mutila. However, when conidia of N. australe and N. ribis were added to the propagation media of potted blueberry plants to investigate the potential for root infection, genotyping by UP-PCR showed that the isolates recovered from the roots and stems of plants were not the isolates added to the soil. Thus, inocula in propagation mixtures is unlikely to cause infection of nursery plants. Isolates of the four main species recovered from farms and nurseries were pathogenic on blueberry stems but pathogenicity differed significantly between species and isolates within a species, with N. ribis being the most pathogenic, then N. parvum, N. luteum and N. australe. Conidia of these species caused necrotic lesions on wounded crowns and trunks, and infection progressed down into roots and up into shoots of the plants. These conidia also caused infection of hard and soft shoots, flower and leaf buds, as well as fruits, incidence and progression usually being greater in wounded than non-wounded tissues. No lesions developed in non-wounded soft and hard shoots. Wound age (0-28 days) in soft and hard shoots and trunks showed decreasing pathogen progression by N. ribis over time, with progression of 16.7, 33.3 and 35.0 mm respectively, by 28 days. However, lesions did not develop from wounds inoculated after 7, 4 and 4 days, respectively. Herbicide damage of hard green shoots allowed for infection and lesion development by N. ribis, levels being similar to the wounded control. The seven blueberry cultivars inoculated with N. parvum and N. ribis conidia differed in disease development with Centra blue, Maru and Ocean blue showing lowest susceptibility. Conidia of the main three Neofusicoccum species were released from pycnidia at 15-30°C and under relative humidities (RHs) of 80-100%, with greatest numbers usually released by N. parvum. Overall, the greatest numbers of oozing pycnidia and conidia released were observed at higher temperatures (25-30°C) and RHs (92-100%). Infection of wounded and non-wounded shoots of potted blueberry plants by N. ribis progressed through the tissues irrespective of the early incubation conditions, 20 or 25°C and 90 or 100% RH. Further, a field study with potted plants showed that N. ribis infection of hard shoots and trunks could take place in summer, autumn and winter, but was least in winter. In vitro fungicide experiments with four isolates each of N. australe, N. luteum, N. parvum and N. ribis showed differences in mycelial growth, conidial germination and germ tube growth between isolates and species, and between products. In vivo evaluation carried out with fungicides on wounded and non-wounded plant tissues on potted and field blueberry plants showed that carbendazim and tebuconazole were the most effective for protecting blueberry plants from infection by Neofusicoccum species. The biological products SerenadeTM and VinevaxTM provided some control if applied 7-14 days before inoculation. Overall, this study has shown that Neofusicoccum species are widespread in New Zealand farms and nurseries and can infect all aerial tissues of the plants, with production of conidia from all tissues. The potential infection of non-wounded tissues has indicated that fungicides need to be applied overall, not just on wounds as previously believed, and that inoculum sources should be eliminated with all possible control methods.