5.2.76
SELECTION OF STREPTOMYCES SPP. WITH POTENTIAL FOR BIOCONTROL OF FUSARIUM OXYSPORUM ON TOMATO

ML SMITHER-KOPPERL and DJ MITCHELL

Plant Pathology Department, University of Florida, Gainesville, FL 32611. USA

Background and objectives
Fusarium wilt (Fusarium oxysporum f.sp. lycopersici, FOL) and Fusarium crown rot (F. oxysporum f.sp. radicis-lycopersici, FORL) cause serious losses in commercial tomato production in Florida [1]. Fumigation and good sanitation practices have been the standard method of control. With the impending loss of methyl bromide as a soil fumigant, and environmental concerns due to the use of pesticides, alternative means of control are being investigated. Various organisms including Trichoderma spp. and Glomus spp. have been evaluated for biological control activity, with some success, but the results tend to be variable. Selected Streptomyces spp. also have the potential to control fungal root and seed rots [2]. The objective of this research was to select Florida isolates of Streptomyces spp. with high potential for biological control activity, based upon their antibiotic activity, capacity to degrade chitin, and lack of harmful effects on the growth of tomato plants.

Materials and methods
Chitinolytic activity: inoculations of 2-mm agar plugs containing mycelium of Streptomyces spp. were made into the center of 6-cm petri plates containing 0.4% chitin agar at pH 8.0. The plates were stored in darkness at 25C and examined after 7 days. The radial distance of clearing in the chitin media around the isolates was recorded and used as an indication of chitinolytic activity. There were three replicates, and the experiments were repeated.
Screening for antibiosis in vitro: Streptomyces spp. were streaked onto cornmeal agar plates in a single line to one side of the middle of the plate. The plates were incubated at 25C under diurnal light. After 7 days, FOL and FORL were inoculated onto the plates on the opposite side from the Streptomyces spp. Hyphal growth of FOL and FORL towards the Streptomyces spp. was measured after 6 days, and compared with growth in the absence of Streptomyces spp. There were three replicates.
Phytotoxic or pathogenic activity: for spore production, Streptomyces spp. were streaked onto YCZ/3 media (113 Difco Czapek's DOX broth, 0.5 g yeast extract, and 20 g agar per liter) in 6-cm petri plates, and incubated in darkness at 25C for 10 days. Roots of 2-week-old tomato seedlings were immersed in spore suspensions prepared in sterile tap water, before transplanting in flats of potting mix in a greenhouse. The experiments were organized in a randomized block design with a treatment unit of five tomato seedlings and five replicates. Survival and phytotoxic symptoms were recorded after 2 weeks.

Results and conclusions
Chitinolytic activity: activity varied between isolates as indicated by the differences in the size of radial clearing in chitin agar. Recorded values for radial clearing range from 0 to 6 mm. A total of 131 isolates with an ability to grow on chitin agar, and with a variable degree of chitinolytic activity, were selected for further trials.
Screening for antibiosis: of the 131 isolates screened for antibiosis, 18 were found to inhibit hyphal growth by 50% or more. In most cases the inhibition was only apparent in hyphal growth towards the Streptomyces isolate; growth away from the isolate was similar to the control. In some isolates the growth of the entire colony was inhibited. One isolate sporulated freely inside the plates and the daughter colonies completely suppressed hyphal growth. The three most promising isolates of Streptomyces spp. were selected on the basis of antibiosis and chitinolytic activity for further evaluations of biological control potential.
Phytotoxic or pathogenic activity: all of the tomato seedlings survived inoculation with the Streptomyces isolates, and there was no evidence of phytotoxic or pathogenic effects after the 2-week trials.

References
1. Kucharek T, Jones JP, Hopkins D, Strandberg J, 1992. Fl. Coop. Ext. Serv./IFAS 1025.
2.Yuan WM, Crawford DL, 1995. Appl. Envir. Microbiol. 61, 3119-3128.