S. C. Kirkpatrick

Fusarium Wilt of Strawberry Caused by Fusarium oxysporum in California

Beginning in 2006 and continuing into 2009, an apparently new disease of strawberry (Fragaria × ananassa) affected commercial plantings (cvs. Albion, Camarosa, and others) in coastal (Ventura and Santa Barbara counties) California. Symptoms consisted of wilting of foliage, drying and withering of older leaves, stunting of plants, and reduced fruit production. Plants eventually collapsed and died. Internal vascular and cortical tissues of plant crowns showed a brown-to-orange-brown discoloration. Differences in cultivar susceptibility were not recorded. Internal crown and petiole tissues, when placed on acidified corn meal agar, consistently yielded Fusarium isolates having similar colony morphologies. No other pathogens were isolated. The Fusarium isolates were subcultured on carnation leaf agar and observed to be producing macroconidia and microconidiophores that are diagnostic of Fusarium oxysporum (1). For two of these isolates, the internal transcribed spacer region comprising ITS1, ITS2, and 5.8S rRNA was amplified using primers ITS-1 and ITS-4 (3). On the basis of a comparison of 515 bp, both isolates had the identical sequence, which was a 100% match for 30 accessions of F. oxysporum in GenBank. This comparison included several formae speciales of F. oxysporum, but F. oxysporum f. sp. fragariae, a previously described pathogen of strawberry (4), was not included. The isolates are archived in the Department of Plant Pathology at UC Davis and are available on request. Both sequenced isolates plus four others were tested for pathogenicity on strawberries. For these tests, spore suspensions of 1 × 105 conidia/ml were prepared separately for six isolates. Roots of strawberry transplants (12 plants of cv. Camino Real) were cut and soaked in spore suspensions for 10 min. Plants were potted in soilless, peat moss-based medium in containers. Control strawberry plants were soaked in water prior to planting. All plants were then grown in a shadehouse. After 8 weeks, inoculated plants began to show wilting and decline of foliage and internal crown tissue was lightly discolored. F. oxysporum was isolated from all inoculated plants. Control plants did not exhibit any disease symptoms and crown tissue was symptomless. To our knowledge, this is the first report of Fusarium wilt of strawberry in California. This disease has been reported from a number of other countries including Argentina, Australia, China, South Korea, Spain, and Japan (2). Since 2006, Fusarium wilt of strawberry has increased in incidence and severity in California. Initial problems in 2006 consisted of multiple small patches (2 to 4 beds wide × 3 to 10 m long) of diseased plants; in these patches disease incidence could range from 80 to 100%. By 2009, in some fields, the disease affected large sections that ran the length of the field. References: (1) P. E. Nelson et al. Fusarium Species: An Illustrated Manual for Identification. Pennsylvania State University Press, University Park, 1983. (2) H. S. Okamoto et al. Plant Prot. 24:231, 1970. (3) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Application. Academic Press, NY, 1993. (4) B. L. Winks and Y. N. Williams. Qld. J. Agric. Anim. Sci. 22:475, 1966.

Limiting Effects of Low Temperature on Growth and Spore Germination in Gibberella circinata, the Cause of Pitch Canker in Pine Species

Pitch canker, caused by Gibberella circinata (anamorph = Fusarium circinatum), causes canopy dieback and mortality in susceptible pine species in many parts of the world. Pitch canker is most problematic in areas with a relatively warm climate, suggesting a possible limitation on disease development imposed by low temperatures. To test this hypothesis, the effect of temperature on radial growth was examined in isolates of G. circinata of diverse geographic origin. All isolates grew most rapidly at 25°C and progressively more slowly at 20, 15, and 10°C. Spore germination occurred most rapidly at 20°C and was slowest at 10°C. To determine if the time required for spore germination might influence the likelihood of infection, the duration of wound susceptibility was examined by inoculating branches of susceptible Monterey pines (Pinus radiata). In each of six field trials, branches were wounded and then inoculated immediately or at 2, 6, or 9 days after wounding. The results indicated that wounds inoculated immediately became infected at a significantly higher rate than those inoculated 2 days later. Thus, if low temperatures extend the time required for germination beyond this period, a reduced infection frequency would be expected. Such a limiting effect of temperature could help to explain the current distribution of pitch canker.

Evidence for a Hemibiotrophic Association of the Pitch Canker Pathogen Fusarium circinatum with Pinus radiata

Fusarium circinatum can be a cause of mortality in pine seedlings but it is also possible for infected seedlings to remain symptomless. The results of this study documented a biotrophic phase in symptomless Pinus radiata seedlings that can persist for at least 52 weeks. A strain of F. circinatum, transformed to express the green fluorescent protein, was observed to grow intercellularly in the root cortex, with no evidence of damage to surrounding cells. Under experimental conditions, shoot symptoms developed only following collar infection, and root deterioration was seen only in plants that first expressed aboveground symptoms. This sequence of events implies that damage to the root system was a secondary consequence of girdling. If so, root symptoms may not reliably detect seedlings infected by F. circinatum. Supplemental mineral nutrition increased the incidence of infection and symptom development in seedlings but some infected plants remained symptomless, precluding the use of this approach to detect infected seedlings. Overall, our findings suggest that the ecological activities of F. circinatum may not be limited to a necrotrophic association with pine trees. A more comprehensive understanding of the life history of this fungus may yield insights that contribute to more effective management of pitch canker.

The Population of Fusarium oxysporum f. sp. fragariae, Cause of Fusarium Wilt of Strawberry, in California

The objectives of this study were to investigate the structure of the population of Fusarium oxysporum f. sp. fragariae in California and to evaluate methods for its detection. Fifty-nine isolates of F. oxysporum f. sp. fragariae were obtained from diseased strawberry plants and their identity was confirmed by pathogenicity testing. The full nuclear ribosomal intergenic spacer (IGS) and elongation factor 1-α gene (EF-1α) were amplified by polymerase chain reaction (PCR) and sequenced to elucidate phylogenetic relationships among isolates. IGS and EF-1α sequences revealed three main lineages, which corresponded to three somatic compatibility groups. Primers designed to detect F. oxysporum f. sp. fragariae in Japan amplified a 239-bp product from 55 of 59 California isolates of F. oxysporum f. sp. fragariae and from no nonpathogenic isolates of F. oxysporum. The sequence of this PCR product was identical to the sequence obtained from F. oxysporum f. sp. fragariae isolates in Japan. Intensive sampling at two locations in California showed results of tests based on PCR and somatic compatibility to be in agreement for 97% (257 of 264) of isolates tested. Our findings revealed considerable diversity in the California population of F. oxysporum f. sp. fragariae, and indications that horizontal gene transfer may have occurred.

Options for Management of Fusarium Wilt of Strawberry in California

ABSTRACT The emergence of Fusarium wilt, caused by Fusarium oxysporum f. sp. fragariae, as a problem for strawberry production in California has been associated with delivery of fumigants to beds through drip-lines rather than flat-fumigation of an entire field. Our research shows that bed fumigation fails to eliminate propagules of the Fusarium wilt pathogen, which remain particularly abundant in bed shoulders at a depth of 30 cm. Controlled environment studies confirmed that inoculum at this depth could cause disease. Screening of currently grown strawberry cultivars documented a wide range of susceptibilities to Fusarium wilt, with some being highly resistant.

Fusarium oxysporum f. sp. mori, a New Forma Specialis Causing Fusarium Wilt of Blackberry

Fusarium oxysporum has recently been identified as the cause of a wilt disease affecting blackberry in California and Mexico. Thirty-six isolates of F. oxysporum obtained from symptomatic blackberry plants in California and Mexico were comprised of nine distinct somatic compatibility groups (SCGs). Phylogenetic analysis of a concatenated data set, consisting of sequences of the translation elongation factor 1-α and β-tubulin genes and the intergenic spacer of the ribosomal DNA, identified nine three-locus sequence types, each of which corresponded to an SCG. Six SCGs were present only in California, two only in Mexico, and one in both California and Mexico. An isolate associated with the most common SCG in California was tested for pathogenicity on blueberry, raspberry, strawberry, and lettuce. All blueberry, raspberry, and lettuce plants that were inoculated remained healthy, but two of the five strawberry cultivars tested developed symptoms. The three strawberry cultivars that were resistant to the blackberry pathogen were also resistant to F. oxysporum f. sp. fragariae, the cause of Fusarium wilt of strawberry. We propose to designate strains of F. oxysporum that are pathogenic to blackberry as Fusarium oxysporum f. sp. mori forma specialis nov.