December 2016 The objectives of this proposal are 1) To determine the temperature and relative humidity optima for Guignardia citricarpa pycnidiospore infection and production on citrus twigs, leaf litter, and fruit; 2) To determine the relative potential of Guignardia citricarpa to form pycnidiospores on citrus twigs, leaf litter, and fruit; 3) To determine whether Guignardia citricarpa can survive and reproduce on citrus debris on grove equipment. Samples collection for inoculum potential continue to be gathered biweekly. Bark is being removed from the twigs and DNA extracted from that. Since determining the best way to extract DNA from the field twigs took longer than expected, processing previously collected twigs continues and qPCR has not been conducted in a systematic manner yet. To get a larger range on temperatures and relative humidities, preliminary studies on the stability of RH measurements for different super saturated salt solutions at a temperature range has been conducted and a statistical analysis to determine whether the variability in RH measurements is excessive for the various salts. The selection will be based on the statistical outcome Experiments were started to look at the effect of temperature on the level of sporulation of P. citricarpa. It can be quite difficult to get consistent sporulation even under controlled conditions. The temperatures that are being tested 15, 20, 24, 28, 32, and 36C. After incubation in complete darkness to avoid the confounding effects of light, it was found for 5 isolates that 24C was the best temperature for sporulation (P < 0.05) followed by 28C. The repetition of the experiment is not yet completed. Work on the effect of FDACS recommended disinfectants (200 ppm bleach or 2000 ppm quaternary ammonium) on conidia germination was conducted. Effective concentrations to inhibit either 50% or 90% of conidia germination for 2 quat products, Canker Solve and C-Quat, and bleach were found to be well below 5 ppm for all products. Bleach was about ten times more effective but is not as stable as quat. The disinfectants have been preliminarily evaluated in the presence of finely ground plant debris (twigs and leaves as would be found on mowers or hedgers). Citrus debris itself had no significant effect on conidia germination but there was a significant effect on the efficacy of the disinfectants. Disinfectant treatment in the presence of citrus tissue debris has a much lower efficacy than determined from previous experiments lacking citrus tissue debris. This loss of efficacy can be attributed to two factors. The first is a reduction in potency due to the presence of tissue debris within the liquid treatment. The second and more profound factor is the availability of disinfectant as a free liquid. When disinfectant is applied at low volumes and entirely or mostly absorbed by the tissue debris, the ai concentration must be raised greater than 200 fold to provide comparable efficacy observed in the presence of free liquid. Confirmatory experiments are being conducted.