Early brown rot infections in sweet cherry fruit are detected by Monilinia-specific DNA primers

Visible and nonvisible quiescent infections of immature and mature fruit ar e an integral component of the disease cycle of brown rot of sweet cherry i n California. Detection of these infections is critical for developing effi cient and efficacious fungicide management programs. The previously publish ed DNA amplification primers mfs3 and NS5 for the identification of Monilin ia fructicola were very specific in amplifying DNA of M. fructicola only an d not M. lawn. This primer set, however, only detected DNA from some of the California isolates of M. fructicola. This genetic diversity was supported by random amplified polymorphic DNA (RAPD) analysis. Using eight 10-mer pr imers, seven M. fructicola isolates from California were all identified as genetically distinct. Using the same primers, only one polymorphism was det ected among seven isolates of M. laxa. The multiple genotypes identified wi thin the small population sample of M. fructicola, but not of M. laxa, usin g RAPD analysis could be indicative of genetic recombination within M. fruc ticola but not within M. laxa. To detect early brown rot infections in frui t, two primer sets that were developed from DNA sequences of either ribosom al DNA (MF5/ITS4/ITS3) or a RAPD fragment (X-09intF3/X-09R) specifically am plified DNA from isolates of M. fructicola and Monilinia species, respectiv ely. No amplification products were present when using DNA from Botrytis ci nerea or from other fungi commonly found on sweet cherry fruit. Primers X-0 9intF3 and X-09R were more sensitive and reliable for detecting small amoun ts of target DNA either extracted from conidia or from laboratory-inoculate d cherry fruit with early brown rot infections that showed no visual sympto ms or with visible quiescent infections. Furthermore, these primers also we re effective for detecting visible quiescent infections in cherry fruit tha t were collected in the field.