Determinação do estado de células viáveis e não cultiváveis e caracterização fisiopatológica de isolados de Xanthomonas citri pv. viticola sob estresse térmico

Abstract

The Northeast region stands out as the largest producer of fine table grapes, corresponding to 99% of Brazilian exports of the fruit. In this region, bacterial canker of the grapevine, caused by Xanthomonas citri pv. viticola, is one of the main plant bacteriosis that threatens this crop. To achieve effective disease management, it is essential to understand the pathogenesis mechanisms of the bacterium, the plant-pathogen interaction, and how the environment affects the pathogen and the disease. In this context, understanding the dynamics of bacterial growth and the influence of temperature on the physio-pathological behavior of bacteria becomes essential to prevent the spread of the pathogen and create efficient strategies for managing the disease. Thus, the present work evaluated the ability to form viable but non-culturable cells (VBNC) and the physio-pathological behavior of X. citri pv. viticola under temperature stress. The CCRMXCV17, CCRMXCV80 and CCRMXCV117 strains had their growth curves determined, the logarithmic phase being observed at 96 h for all strains, while the death phase was observed at 480 h for the CCRMXCV17 strain and 288 h for the other two strains. With qPCR analysis, the occurrence of VBNC state was confirmed at 480 h for the strain CCRMXCV17 and 288 h for CCRMXCV80 and CCRMXCV117. The incubation period (IP) and the final severity (FS) of the artificial inoculation of the strains in the logarithmic and VBNC phases, did not obtain a significant interaction (P>0.05) between the phase and isolate factors, however, there was a significant difference in the IP between the means of the strain factor. All strains were resuscitated at the VBNC stage after inoculation in NYD medium. However, only the strain CCRMXCV117 showed a significant difference between the concentrations of the exponential and VBNC phases. The strains were cultivated at temperatures 29, 33, 37, 41, and 45 ºC and inoculated in grapevine seedlings to determine the epidemiological components of bacterial canker and were evaluated for in vitro biofilm formation and influence of swarming motility. The inoculation of strains cultivated under temperature stress showed a significant interaction between the factors, strain and temperature, only for the FS parameter. Weak biofilm matrix formation was observed in all strains, with a significant interaction between the factors, temperature and strain. Swarming motility under temperature stress was not detected in all three stains, since the colony growth was restricted to the center of the plate, however, there was a significant difference between the means of the factors evaluated individually. This is the first report of the occurrence of VBNC condition in strains of X. citri pv. viticola, as well as the pathogenic capacity at this stage. Thus, we emphasize the importance of applying a set of techniques for more accurate detection of this disease, as well as the continuous research of other characteristics, beyond biofilm and motility, that may influence the mechanisms of pathogenicity.