THE ISOLATION AND IDENTIFICATION O F SEVERAL TRICHOTHECENE MYCOTOXINS FROM FUSARIUM HETEROSPORUM R. J. COLE*and J. W. DORNER Mational Peanut Research Laboratory, U S D A ,1 S E A , A R , S R , P . 0 . Box 637, Dawson, Georgia 317.42
R. H. Cox National Institute of Environmental Health Sciences, P . 0. Box 18833, Research Triangle Park, N . C . 87709
B. M. CUXFER Georgia Experiment Station, Experiment, Georgia 3081 8
H. G. CUTLER U S D A , S E A , A R , S R , Georgia Coastal Plain Experiment Station, Tqton, Georgia 31 795 and
B. P. STUART Veterinary Diagnostic and Investigational Laboratory, University of Georgia, Tifton, Georgia 31794
ABSTRACT-^ isolate of Fusarium heterosporum Nees ex Fr. that parasitizes the honeydew and immature sclerotia of Claviceps paspali was shown to be highly toxigenic to day-old chickens. The major toxicity of culture extracts of the fungus was due to the presence of several trichothecene mycotoxins. Six trichothecenes were isolated and chemically identified by spectroscopic methods as T-2 toxin (l),HT-2 toxin (2) , T-2 tetraol ( 6 ) ,3a,4~,dihydroxy-15acetoxy-8a-(3-hydroxy-3-methylbutyryloxy)-l2,13-epoxytrichothec-Q-ene (3), 3a,4~,15trihydroxy&-(3-hydroxy-3-methy1butyryloxy)-12,13-epoxytrichothec-9-ene( 5 ) , and 3a,48,~-trihydroxy-l5acetoxy-l2,13epoxytrichothec-sene (4). The acute toxicities of the trichothecenes were compared t o the effects of T-2 toxin in day-old chickens and on the back skins of rabbits. The comparative inhibitory effects of the trichothecene mycotoxins in the Triticum coleoptile bioassays were also determined.
Fusarium heterosporum Nees ex Fr. is an ubiquitous saprobic fungus that colonizes the honeydew and immature sclerotia of Claoiceps paspali, C . purpurea, and probably most other ergot fungi (1,2,3). As F . heterosporum colonizes and overgrows the honeydew, it prevents the maturation of the sclerotium and thus acts as a natural biological control. Matured sclerotia of C . purpurea that were partially colonized by F . heterosporum rotted rapidly and did not germinate when moistened (Cunfer and Mathre, unpublished). Colonization by F . heterosporum is easily identified because it produces a bright pink to orange mycelial stroma over most of the immature sclerotium. The stroma is composed of dense masses of multiseptate conidia. These conidia are disseminated in the same manner as ergot conidia. When F . heterosporum and ergot conidia reach a susceptible flower at the same time, F . heterosporum does not prevent infection by the ergot fungus. However, as soon as honeydew appears, the F . heterosporum conidia germinate and colonize the honeydew (1). *To whom correspondence should be addressed. 'Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the U.S. Department of Agriculture and does not imply its approval to the exclusion of other products that may also be suitable.
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MAY-JUN
19811
COLE ET AL. :
TRICHOTHECENE MYCOTOXINS
325
By the end of each summer in Georgia and throughout most of the southern United States, 30 to 50% of C. paspali stromata infecting dallisgrass (Paspalum diZafatum Poir.) flowers are colonized by F . heterosporum. F . heterosporum produces abundant numbers of conidia, both in vitro and in vivo, and chlamydospores that persist for long periods until they reach a favorable substrate. Several taxonomic systems are used for the Fusaria. F . heterosporum is the species name according to the scheme of Booth (4). In the Snyder and Hansen (5) system, it is F . roseum Lk. ex Fr.; according to the Woolenweber (6) system, it is called F . sambucinum Fuckel. The potential for mycotoxin production by F . heterosporum on ergot honeydew has been previously studied. Mower et al. ( 3 ) were unable to detect psychotrophic compounds from cultures of F . heterosporum isolated from C. purpurea or from cultures of F . heterosporum grown on ergotamine tartrate.
1 R1 = OAc, Re 2 R1 -
= OH,
=
OAc, R3
R2 = OAc, R
= OCOCH2CH(CH3)2
3 = OCOCH2CH(CH3)2
3 R1 = OH,
Re = OAc, R3 = OCOCH2COH(CH3)2
4 R, = OH,
R2 = OAc, R j = OH
5 R1
= OH,
R2 = OH,
R3
= OCOCH2COH(CH3)2
6 R1
= OH,
R2 = OH,
R3
= OH
FIG.1. Chemical structures of some trichothecene my cot oxins.
We became interested in the toxin-producing potential of F . because of its widespread occurrence in nature. We now report and identification of several trichothecene mycotoxins from F . isolated from C. paspali sclerotik in middle Georgia in the summer of parative acute toxicities of the trichothecenes are presented.
heterosporum the isolation heterosporum 1977. Com-
EXPERIMEXTAL ISOLATION A N D CULTURE OF FUh'Gm-The strain of Fusarium heterosporum Nees ex Fr. [= F. roseum Lk. ex Fr.] was isolated from Claziceps paspali honeydew on Paspalum distichum L. a t Experiment, Ga., in the summer of 1977. The isolate was grown on fresh potato-dextrose agar a t 20 to 25" in light supplied by fluorescent tubes. Under these conditions, F. hetero-
mm
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- $OH -HT-2 15Acatoxy T-2 3: OH -T-2 Triol ------------ -.----. - ---..----------- -Tctraol
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MOLAR CONCENT RAT ION FIG.2. Inhibitory effects on the growth of wheat coleoptiles (Triticum aestivum L. cv. Wakeland) by some trichothecene mycotoxins. Control, dotted line. Significant inhibition below solid line (P
