Visual markers detectable by histochemical staining have been developed
for analysing the time course and tissue specificity of maize
infections by Fusarium moniliforme. Three F. moniliforme
strains, RRC 374, MRC 826 and RRC PAT, were transformed with a
plasmid, pHPG, containing the gusA reporter gene which codes for
β-glucuronidase (GUS) and the hph gene for hygromycin
resistance as the selectable marker. Introduction of plasmid DNA into germinating
conidia yielded 1·2×10−7
transformants per conidium; expression of both gusA and hph
was however, transient. Stable transformants were obtained using protoplasts
as the
recipient, but transformation frequency was reduced. Southern blot and
PCR analyses confirmed incorporation of pHPG into the
genome of all three F. moniliforme strains with gusA
properly inserted in MRC 826 and RRC PAT, but apparently disrupted in RRC
374. The growth pattern for transformed F. moniliforme isolates
and the parental wild types followed a sigmoid curve on minimal
and enriched media. Hygromycin totally inhibited growth for wild type isolates,
but not of transformants. Transformed isolates
maintained the ability to infect the maize plant. Thus, this study is the
first report of F. moniliforme transformed with a visibly
detectable reporter gene to use for analysing this endophyte-host interaction
of world-wide importance to animal and human health.