First Report of Rust Caused by Puccinia emaculata on Switchgrass in Arkansas.

In May 2007, switchgrass (Panicum virgatum L.) cv. Alamo and a breeding line, OSU-NSL 2001-1, were planted at the Arkansas Agricultural Research and Extension Center, Fayetteville. In August 2008, a high incidence of dark brown-to-black rectangular foliar lesions delineated by major veins was observed throughout plots of both lines. Lesions covered 25% to nearly 100% of total leaf tissue. Similar symptoms were also observed on unknown switchgrass cultivars in Benton County in northwest Arkansas and in St. Francis County in east-central Arkansas, suggesting that the disease was widely distributed throughout the state. The pathogen produced epiphyllous and adaxial masses of dark brown-to-black telia from erumpent fissures on leaf surfaces. Dark brown teliospores were observed under magnification and were two-celled, oblong to ellipsoid, and 33 ± 3.5 µm long with an apical cell width of 17.5 ± 2.7 µm and basal cell width of 16.2 ± 2.8 µm (reported as mean ± standard deviation, n = 25). Pedicles were colorless to light brown and measured 25.4 ± 9.2 µm (n = 25). In June 2009, at the Fayetteville Research and Extension Center, several second-year stands of switchgrass developed amphigenous and adaxial foliar lesions containing urediniospores. The uredia were globose and finely echinulate, measuring 23.1 ± 2.2 µm (n = 25) with brown cell walls. Teliospore and urediniospore morphology from all collections was consistent with Puccinia emaculata Schw. (2). Genomic DNA was extracted from a representative infected leaf of cv. Alamo, collected in Fayetteville, AR in June 2009, and amplified by PCR with primer sets PRITS1F (3) and ITS4B (1), which amplified an 803-bp fragment of rDNA encoding the first internal transcribed spacer (ITS1), 5.8S subunit, and second internal transcribed spacer (ITS2). The fragment was cloned into pGEM T Easy (Promega Corp, Madison, WI) and sequenced. A BLAST search of GenBank revealed that the fragment was most similar to the rDNA of P. emaculata (GenBank Accession No. EU915294.1; 755 of 758 bases matching; 99% identity) previously reported as a pathogen on switchgrass in Tennessee (3). The incidence and severity of rust on the widely planted switchgrass cv. Alamo is considerable cause for concern as efforts are made to increase acreage and production. Climatic conditions in St. Francis County are generally consistent with locations in Tennessee where switchgrass rust was previously reported (3). However, northwest Arkansas represents the eastern edge of the southwestern United States, suggesting that P. emaculata may affect switchgrass in geographically diverse areas of the United States. To our knowledge, this study represents the first report of rust on switchgrass in Arkansas. Managing this disease will be an important consideration for large-scale switchgrass cultivation in the state. References: (1) M. Gardes and T. D. Bruns. Mol. Ecol. 2:113, 1993. (2) P. Ramachar and G. Cummins. Mycopathol. Mycol. Appl. 25:7, 1965. (3) J. Zale et al. Plant. Dis. 92:1710, 2008.

Development of Anthracnose on Grain Sorghum Hybrids Inoculated with Recently Described Pathotypes of Colletotrichum sublineolum Found in Arkansas.

Sorghum anthracnose, caused by Colletotrichum sublineolum, is found in most grain-sorghum-producing areas, including Arkansas. Yield losses can be severe in susceptible sorghum hybrids, suggesting that host resistance will continue to be critical for maintaining high yields. However, genetic control is often unsuccessful because of haplotype and pathotype variability within the C. sublineolum population. The objective of this research was to assess the levels of resistance of selected grain sorghum hybrids to isolates of the recently described pathotypes of C. sublineolum found in Arkansas. Field experiments were conducted in 2005 and 2006 to evaluate the reaction of 10 sorghum hybrids to eight C. sublineolum pathotypes by collecting anthracnose severity ratings at 7, 14, 21, and 28 days after inoculation. Based on these disease severity ratings, the area under the disease progress curve was calculated for each pathotype by hybrid interaction. These data showed that different levels of resistance are available in the current grain sorghum hybrids to different pathotypes. Cargill 888Y was resistant to all pathotypes in both years. However, Asgrow A571, DEKALB DKS53-11, FFR 318, and Pioneer 84G62 were moderately susceptible to susceptible to all pathotypes tested in both years. Pathotype 9 caused the most disease on 7 of the 10 hybrids evaluated in 2005 but, in 2006, pathotypes 12 and 13 surpassed pathotype 9 for disease severity on 9 of the 10 hybrids. More work is needed to assess host resistance and the effect of sorghum anthracnose on yield parameters of grain sorghum hybrids planted in Arkansas.

Pathotypes of Colletotrichum sublineolum in Arkansas.

Sorghum anthracnose, caused by Colletotrichum sublineolum, is widely distributed in most grain sorghum-producing areas in Arkansas, and localized epidemics continue to occur across the state. The objectives of this research were to determine the number of pathotypes of C. sublineolum in Arkansas and to determine where these pathotypes are located within the state. Ninety-eight isolates were collected from research stations and production fields located in the Delta region of Arkansas from 2003 through 2005 and were tested for virulence to eight differential grain sorghum breeding lines. Of these, 11 isolates produced conflicting and unreliable results and were excluded from further analysis. Among the remaining 87 isolates, 13 pathotypes were identified. Eleven of these pathotypes were previously unreported, with 53% of the isolates consisting of pathotype 9. Pathotype 9 was distinguished from all other pathotypes by its virulence to all eight differential lines. Pathotype diversity appeared to be more closely associated with the year than with the locations where isolates were collected. Although all host differential lines were susceptible to at least 50% of the isolates evaluated, inbred lines SC326-6, TAM428, and TX2536 were susceptible to more than 90% of the 87 isolates evaluated. These results showed that the C. sublineolum populations in Arkansas are diverse based on the comparisons of pathogenicity to previously published host differentials, and it appears that these field populations varied from year to year.

Infection of Rice Seed Grown in Arkansas by Pyricularia grisea and Transmission to Seedlings in the Field.

Rice blast, caused by Pyricularia grisea, is an important and serious disease of rice (Oryza sativa) in the southeastern United States. The disease sporadically reaches epidemic proportions on susceptible cultivars within fields and over large areas within Arkansas. The main overwintering sources of inoculum reportedly include infected rice stubble, related host species, and infected seed. The objectives of the research were to (i) determine whether rice seed grown in Arkansas were infected with P. grisea, (ii) investigate the relationship between seed infection and seedling disease, and (iii) determine if planting naturally infected seed could lead to the subsequent development of rice blast on seedlings in the field. The results of seed assays showed that P. grisea was detected in samples of foundation, certified, and production seed. Estimated levels of infection by P. grisea of rice seed from 66 samples of rice seed grown in Arkansas ranged from 0 to 10.5%. Planting infected seed in the greenhouse and the field resulted in seedling infection. Planting naturally infected seed may result in disease development (i) from seedlings grown from infected seed planted beneath the soil surface, (ii) from seedlings grown from germinating seed left on the soil surface, (iii) from seed coats, or (iv) from nongerminated seed left on the soil surface after planting. Additional research is necessary to establish the mechanisms of infection of seedlings and to establish disease thresholds for this important fungal pathogen of rice.

Rice Blast Epidemics Initiated by Infested Rice Grain on the Soil Surface.

Field experiments were conducted in 1996 and 1997 with a marked strain of Pyricularia grisea to determine if inoculum from infested rice grain could cause primary infections and sustain a rice blast epidemic during the growing season by giving rise to leaf, collar, and neck symptoms. The marked strain, a sulfate nonutilizing (sul) mutant of P. grisea, was grown on autoclaved rice seed for 7 days at 25°C. Infested rice grains were applied to the soil surface at the time of plant emergence (approximately 10 days after planting) at densities of 0, 0.5, 5, 25, and 50 grains per 0.1 m2 in plots planted to the blast susceptible cv. M-201. Leaf blast symptoms were first detected in the plots containing infested grain 35 days after plant emergence in both 1996 and 1997. The sul mutant was isolated from more than 90% of the lesions sampled from rice seedlings 35 to 45 days after plant emergence. Leaf blast increased more rapidly in plots with 25 and 50 infested grains per 0.1 m2 than in plots with less inoculum pressure (0.5 and 25 infested grains per 0.1 m2), although in 1996, leaf blast incidence recorded at midseason in plots containing 0.5 and 5 infested grains per 0.1 m2 was 41 and 55%, respectively. At the end of both seasons, the sul mutant was recovered from over 90% of the leaf, collar, and neck blast lesions except for one sample date in 1996. Rice blast was not detected in the control plots (no infested grain) in 1997 and not until 65 days after planting in 1996. Comparisons of disease progress on leaves between the marked strain and the parental wild-type strain under field conditions indicated that development of disease caused by the sul mutant was similar to disease caused by the wild-type strain.

Effect of Nitrogen Fertilization on Disease Progress of Rice Blast on Susceptible and Resistant Cultivars.

The effects of three nitrogen fertilization treatments on the development of rice blast were studied on eight cultivars under field conditions in Arkansas in 1995 and 1996. The eight cultivars (Kaybonnet, Cypress, Lacassine, Mars, Adair, Alan, Newbonnet, and RT7015) ranged from resistant to susceptible to blast according to previous field observations. The recommended nitrogen levels for the eight cultivars varied from 123 to 168 kg/ha/year. Three treatments, consisting of different rates and timing of nitrogen applications, were tested over 2 years at one location. The first treatment consisted of a single nitrogen (N) application applied to plots at the recommended rate at preflood during the midtillering stage. The second treatment consisted of applying nitrogen as a single preflood application but at 1.5 times the recommended N rate used in treatment one. The third treatment (control) consisted of applying the recommended amount of nitrogen fertilizer used in treatment one, but in a three-way-split application with 56 to 100 kg/ha (depending on the cultivar) of urea applied at preflood followed by the application of 34 kg/ha of N applied approximately 10 and 20 days after the panicle differentiation (PD) growth stage. Inoculated spreader plots were used to initiate rice blast epidemics in the test plots. The results indicate that the disease progress for rice blast, regardless of N treatments, followed a unimodal curve, whereby disease incidence and total lesion area per plant reached a maximum near midseason (PD growth stage) and then gradually declined. This decline in disease was attributed to adult resistance, leaf senescence, and the formation of new (noninfected) leaves. Application of nitrogen above the recommended rate for any given cultivar significantly increased disease incidence and total lesion area per plant on all cultivars except Kaybonnet, a highly resistant cultivar. Furthermore, a differential cultivar response to nitrogen was observed when measuring both disease incidence and total lesion area per plant. Leaf blast was significantly more severe on the susceptible and very susceptible cultivars when N fertilizer was applied as a single application at preflood than in the split application treatment. Nitrogen treatments did not significantly affect the incidence of collar rot or neck blast.

Mating system of the filamentous ascomycete, Glomerella cingulata.

Mating in heterothallic filamentous ascomycetes is typically controlled by a single mating-type locus with two alternate alleles or idiomorphs. In this study, five self-sterile strains of Glomerella cingulata from pecan were crossed in all possible combinations. Four of the five strains could be placed into two mating-type groups, but the fifth strain was sexually compatible with all of the other strains. Single ascospore progeny were isolated from each of the successful crosses, tested for self-fertility, and backcrossed with both parents. In addition, subsets of F1 isolates were crossed with all five of the original strains from pecan and in all possible combinations with each other. Results from the crosses showed that the ascospore progeny had stably inherited the mating pattern of one of the parental strains and that the mating type had segregated 1:1 among the F1 isolates. Furthermore, the five strains from pecan were sexually compatible with five additional heterothallic strains in all but one combination. Data from these experiments are consistent with a mating system composed of a single mating-type locus with multiple alternate alleles. We believe that this is the first report of this type of mating system for an ascomycete species.

Behavior of a Wild-Type and Two Mutant Strains of Colletotrichum gloeosporioides f. sp. aeschynomene on Northern Jointvetch in the Field.

The fungus Colletotrichum gloeosporioides f. sp. aeschynomene causes an anthracnose on Aeschynomene virginica and has been used as a biological control agent to control this weed in the United States. The population dynamics of a wild-type strain (3-1-3) and two mutant strains of 3-1-3 of C. gloeosporioides f. sp. aeschynomene, a benomyl-resistant strain (B21) and nitrate-nonutilizing strain (Nit A), were studied in field tests on northern jointvetch in 1994 and 1995 to determine how the strains interacted on infected plants under field conditions. Plants were co-inoculated with strains 3-1-3 and B21, strains 3-1-3 and Nit A, and strains 3-1-3, B21, and Nit A at equal and unequal initial proportions. Plants were grown and maintained under flooded conditions in small wading pools. In co-inoculation of plants with 3-1-3 and B21 from equal initial proportions, the population of 3-1-3 increased slightly until it reached a proportion of 60 to 70%, whereas the population density of B21 reached 30 to 40% at the end of growing season. From unequal initial proportions, the population density of B21 decreased from 90 to about 50%, whereas the 3-1-3 increased from 10 to 50%. The population density of 3-1-3 increased from an equal initial proportion and was significantly greater than that of Nit A on every sampling time. From unequal initial proportions, the population density of 3-1-3 increased from 10 to 90%, whereas that of Nit A declined. In co-inoculation of plants with the three strains, the population density of 3-1-3 was significantly greater than those of the mutant strains at every sampling time. The proportions of mutant strains within the total population of C. gloeosporioides f. sp. aeschynomene on plants varied according to the test conditions and the number and types of strains co-inoculated.

Infection Components of Wild-Type and Mutant Strains of Colletotrichum gloeosporioides f. sp. aeschynomene on Northern Jointvetch.

Colletotrichum gloeosporioides f. sp. aeschynomene causes an anthracnose of northern jointvetch, Aeschynomene virginica. Infection components, including lesion number, latent period, lesion expansion rate, and sporulation, were measured in experiments conducted in controlled environments. Two wild-type strains (3-1-3 and CLA 5A), four benomyl-resistant strains (B13, B15, B18 and B21), and four nitrate nonutilizing mutant strains (Nit A, Nit R, Nit L, and Nit T) of the pathogen were tested. Nitrate nonutilizing strains caused significantly fewer lesions on northern jointvetch than did wild-type and benomyl-resistant strains. Latent periods were significantly shorter for the wild-type strain CLA 5A than for most other strains. Lesion expansion rates of all benomyl-resistant strains were significantly slower than those of the wild- type strains. Large variations in sporulation were observed for most strains, and no differences in sporulation were found between wild-type and mutant strains. The usefulness of infection component analysis for the identification of competitiveness of strains of fungal pathogens for biological control of weeds is discussed.

Evidence for mating between isolates of Colletotrichum gloeosporioides with different host specificities.

Individual isolates of the ubiquitous plant pathogen Colletotrichum gloeosporioides (teleomorph Glomerella cingulata) can have very restricted host ranges. Isolates that share the same host range are considered to be genetically discrete units, and sexual compatibility has been reported to be limited to individuals that share the same host range. However, we have recently observed that some isolates of C. gloeosporioides that are specifically pathogenic to different, distantly-related hosts are sexually compatible. Ascospore progeny from one such cross were randomly isolated and outcrossing was verified by the reassortment of several RFLP markers among the progeny. In addition, the progeny were analyzed for pathogenicity to parental hosts. The implications of sexual compatibility between C. gloeosporioides isolates with different host specificities on the evolution of Colletotrichum species are discussed.