Development of a quantitative polymerase chain reaction detection protocol for Cercospora kikuchii in soybean leaves and its use for documenting latent infection as affected by fungicide applications

Cercospora leaf blight (CLB) of soybean, caused by Cercospora kikuchii, is a serious disease in the southern United States. A sensitive TaqMan probe-based real-time quantitative polymerase chain reaction (qPCR) assay was developed to specifically detect and quantify C. kikuchii in naturally infected soybean plants. The sensitivity was 1 pg of genomic DNA, which was equivalent to about 34 copies of genome of C. kikuchii. Using this qPCR assay, we documented a very long latent infection period for C. kikuchii in soybean leaves beginning at the V3 growth stage (as early as 22 days after planting). The levels of biomass of C. kikuchii remained low until R1, and a rapid increase was detected from the R2/R3 to R4/R5 growth stages shortly before the appearance of symptoms at R6. The efficacy of various fungicide regimens under field conditions also was evaluated over a 3-year period using this qPCR method. Our results showed that multiple fungicide applications beginning at R1 until late reproductive stages suppressed the development of C. kikuchii in leaves and delayed symptom expression. Different fungicide chemistries also had differential effects on the amount of latent infection and symptom expression during late reproductive growth stages.