Genetic Diversity and Fitness of Fusarium graminearum Populations from Rice in Korea

Fusarium graminearum is an important fungal pathogen of cereal crops and produces mycotoxins, such as the trichothecenes nivalenol and deoxynivalenol. This species may be subdivided into a series of genetic lineages or phylogenetic species. We identified strains of F. graminearum from the Republic of Korea to lineage, tested their ability to produce nivalenol and deoxynivalenol, and determined the genetic composition and structure of the populations from which they were recovered. Based on amplified fragment length polymorphism (AFLP), PCR genotyping, and chemical analyses of trichothecenes, all 249 isolates from southern provinces belonged to lineage 6, with 241 having the nivalenol genotype and 8 having the deoxynivalenol genotype. In the eastern Korea province, we recovered 84 lineage 6 isolates with the nivalenol genotype and 23 lineage 7 isolates with the deoxynivalenol genotype. Among 333 lineage 6 isolates, 36% of the AFLP bands were polymorphic, and there were 270 multilocus haplotypes. Genetic identity among populations was high (>0.972), and genotype diversity was low (30 to 58%). To test the adaptation of lineage 6 to rice, conidial mixtures of strains from lineages 3, 6, and 7 were inoculated onto rice plants and then recovered from the rice grains produced. Strains representing lineages 6 and 7 were recovered from inoculated spikelets at similar frequencies that were much higher than those for the strain representing lineage 3. Abundant perithecia were produced on rice straw, and 247 single-ascospore isolates were recovered from 247 perithecia. Perithecia representing lineage 6 (87%) were the most common, followed by those representing lineage 7 (13%), with perithecia representing lineage 3 not detected. These results suggest that F. graminearum lineage 6 may have a host preference for rice and that it may be more fit in a rice agroecosystem than are the other lineages present in Korea.Fusarium graminearum (teleomorph: Gibberella zeae) causes head blight of small grains, including rice, wheat, and barley (23). The fungus was first reported on rice in Italy by Cattaneo (4) as Botryosphaeria saubinetii Niessland. This rice disease has since been recorded in other countries, including Brazil, China, India, Japan, Nepal, and Uganda (11, 31). The disease usually does not cause heavy damage, but under conditions that favor disease development, e.g., high humidity, it may be severe. Chung et al. (7) found that an isolate from wheat could infect rice and other plants and also could cause a postemergence blight in rice. Wheat isolates of F. graminearum can cause significant disease on rice, but under greenhouse conditions no trichothecenes were detected in the infected rice florets (14). In addition, Nepalese rice contained no detectable contamination with trichothecenes even though F. graminearum occurs in Nepal (11).The fungus can produce the 8-ketotrichothecene mycotoxins nivalenol (NIV) and deoxynivalenol (DON). Most of the biosynthetic genes for the synthesis of 8-ketotrichothecenes are tightly linked in the TRI gene cluster (9). TRI7 and TRI13 are required for acetylation and oxygenation of the oxygen at C-4 to produce NIV and 4-acetyl nivalenol (4-ANIV), respectively, from DON. PCR-based methods to identify polymorphisms in both genes were developed as simple, reliable diagnostic tools for differentiating strains with DON and NIV chemotypes (20, 21). There are regional differences in the distribution of the two chemotypes. Maize and wheat in North America and Europe commonly are contaminated with DON (9), while strains with NIV chemotypes are commonly recovered from cereal crops in Asia (15, 17). In the Republic of Korea, strains with the DON chemotype often cause maize ear rot, while strains with the NIV chemotype commonly are recovered from barley (17, 35). A severe epidemic of Fusarium head blight on wheat and barley occurred in 1963 in southern Korea (5, 6). Humans and farm animals consuming moldy cereals exhibited typical signs of trichothecene intoxication involving vomiting, dizziness, nausea, abnormal pain, and diarrhea (9). The natural occurrence of NIV and DON has been reported in barley and maize in Korea (17, 35, 41), but there have been few surveys of Fusarium mycotoxins in Korean rice.O''Donnell et al. (30) divided F. graminearum into seven phylogenetic lineages based on the genealogical concordance of six genes. The phylogenetic separation has been used to raise these seven and four additional lineages to species status (36). The geographic location often influences the lineage present, e.g., lineage 7 is the most common in the United States, and lineage 6 dominates in China. Lineage and trichothecene chemotype are not correlated (45), and the lineages are morphologically cryptic. Members of all lineages are cross-fertile with strains belonging to lineage 7 and in some cases with strains of other lineages (1, 2, 19, 25), a pattern that suggests that the members of all of the lineages belong to a single biological species.Studies of F. graminearum populations have been made in different geographic regions, e.g., China (12), Europe (42), the United States (48, 49), and Argentina (34). Populations of F. graminearum have high levels of genotypic diversity, which suggests that recombination occurs regularly in F. graminearum populations. Most studies have focused on populations from wheat, barley, and corn, and there is little information on F. graminearum populations from rice.Severe epidemics of Fusarium head blight of rice occurred in August 2001 after heavy rainfall during the rice flowering period in southern Korea. Lesions on or discoloration of the glumes were common, with infected grains first appearing to be white and later yellow, salmon, or carmine. Sometimes the entire seed was colonized. Infected grains were lightweight, shrunken, and brittle. Our objectives in the present study were (i) to determine the frequency at which F. graminearum occurs in plants with rice head blight; (ii) to determine the number and relative frequency of the F. graminearum lineages present; and (iii) to evaluate the strains for their sexual fertility, genetic relatedness, virulence, and toxin-producing abilities. Our working hypotheses were that sexually fertile strains from lineage 6 would dominate in the population and that these strains would be the most aggressive toward rice. We expected most of the lineage 6 strains to produce NIV and for there to be a high level of genetic variation, as assessed by neutral (amplified fragment length polymorphism AFLP]) markers. We evaluate here F. graminearum population diversity in Korea and provide new information on the pathogenic capabilities of strains belonging to several of the known lineages of this very widespread fungal species.