Loblolly pine (Pinus taeda) is the major commercial pine species cultivated in the Gulf Coast Region of the southern United States. Symptoms of Diplodia shoot blight (including yellow and brown needles and resin-soaked, dead, small twigs), pycnidia with conidia typical of Diplodia pinea on blighted shoots, and damaged, immature seed cones were observed during the summer of 2007 in loblolly pine seed orchards near Ward, AL, Winn Parish, LA, and Moselle, MS. Similar conidia also were obtained from pycnidia on opened seed cones of longleaf pine (P. palustris) collected on the campus of Mississipi State University, Starkville. Pure cultures obtained from specimens collected at each location were confirmed as D. pinea using species-specific PCR primers (3) that allow differentiation of D. pinea from the similar pine shoot blight pathogen D. scrobiculata. Isolates from loblolly pines in Alabama (07-58), Louisiana (07-38), and Mississippi (06-45) were used individually to inoculate potted 6- to 7-month-old loblolly pine seedlings grown from seed in a greenhouse in each of two independent trials. Elongating terminal shoots of seedlings to be inoculated were wounded by removing a needle fascicle ∼2 cm below the shoot apex. A 4-mm-diameter plug cut from an actively growing colony on water agar (WA) was placed on the wound, mycelium side toward the stem. Noncolonized WA plugs were placed in the same manner on similarly wounded control seedlings and nonwounded control seedlings also were used. Parafilm was wrapped around the shoots to hold the agar plugs in place and was removed after 1 week. Each of the five isolate-treatment combinations was applied to seven (trial 1) or eight (trial 2) seedlings (35 and 40 seedlings per trial, respectively). One week after inoculation, small, brown lesions were visible at the point of inoculation on stems of most of the inoculated seedlings. At 25 days after inoculation, all inoculated seedlings exhibited needle browning and stem cankers ranging from 0.6 cm to 9.0 cm long (mean 2.5 cm) that girdled and killed distal portions of the shoots of ∼25% of the inoculated seedlings in each trial. Wounded control and nonwounded control seedlings did not develop symptoms. Stem segments including the point of inoculation (or comparable segments of wounded and nonwounded control seedlings) were excised, surface disinfested, and incubated on tannic acid agar with sterile red pine needles. D. pinea was cultured from all inoculated seedlings and also from one wounded control seedling. Although occurrence of D. pinea on Cedrus spp. is included in an index (1), to our knowledge this is the first confirmed report of D. pinea on pines in Alabama, Louisiana, and Mississippi. The degree of risk presented by D. pinea to loblolly pine, longleaf pine, and other pine species native to the southern United States when grown in their native ranges is unknown. Reports of Diplodia shoot blight of southern U.S. pines when grown as exotics in the southern hemisphere (4) and the potential for epidemics to develop suddenly under severe weather conditions (2,4) justify additional studies to evaluate the potential for damage to these hosts in their native ranges. References: (1) Anonymous. Page 333 in: Index of Plant Diseases in the United States. Agric. Handb. 165, U.S. Dep. Agric. Washington, DC, 1960. (2) T. H. Nicholls and M. E. Ostry. Plant Dis. 74:54, 1990. (3) D. R. Smith and G. R. Stanosz. Plant Dis. 90:307, 2006. (4) W. J. Swart and M. J. Wingfield. Plant Dis. 75:761, 1991.

Spring dead spot (SDS) is the most destructive disease of bermudagrass (Cynodon dactylon (L.) Pers.). Symptoms of SDS appear in the spring when bermudagrass transitions out of winter dormancy. These symptoms include depressed, straw-colored patches that range from several centimeters to a meter in diameter. Infected roots and rhizomes are black, brittle, and necrotic. The disease is caused by three species of fungi: Ophiosphaerella herpotricha (Fr:Fr) J. Walker; O. korrae (J. Walker & A.M. Smith) Shoemaker & C.E. Babcock; or O. narmari (J. Walker & A.M. Smith) Wetzel, Hubert & Tisserat. However, O. korrae is the most prevalent causal organism of SDS in the southeastern United States and was the only species reported in Mississippi (1). In April of 2006, root samples were collected from a bermudagrass putting green in Booneville, MS with a high level of SDS incidence and severity. Symptomatic roots were collected and surface disinfested in 0.6% NaOCl and plated on one-quarter-strength potato dextrose agar (PDA) amended with streptomycin sulfate and chloramphenicol. Hyphae growing from the roots were transferred to full-strength PDA after 5 to 7 days. Mycelium from five pure-culture colonies plus an O. herpotricha control was harvested after 4 weeks of growth and the genomic DNA was extracted. The genomic DNA of the Booneville isolates and the O. herpotricha control were amplified by PCR using species-specific primers OHITS1 and OHITS2 for O. herpotricha (2). Amplification of a 454-bp fragment of DNA confirmed one of the five unknown isolates as O. herpotricha. The other four isolates were not identified. 'Sahara' bermudagrass (4 weeks old in 3.8 × 20 cm Cone-tainers containing a sand and soil mixture) was inoculated with the Booneville-O. herpotricha isolate and the O. herpotricha control. One gram of oat seed infested with O. herpotricha isolates was inserted 2 cm below the crowns in the root zone of bermudagrass plugs. The inoculated bermudagrass plants were incubated for 4 weeks in the greenhouse. A control consisting of noninfested sterile oats was included. Following incubation, black, necrotic roots were observed on the plants inoculated with both O. herpotricha isolates. No symptoms were observed on roots of noninfested plants. Symptomatic roots were disinfested and plated on one-quarter-strength PDA. Koch's postulates were completed after O. herpotricha was reisolated from roots of plants inoculated with both O. herpotricha isolates and confirmed by PCR as mentioned above. The identification of O. herpotricha as a causal organism of SDS in Mississippi clarifies the involvement of multiple causal agents in this state and broadens the geographic distribution of this root-rot species. References: (1) F. B. Iriarte et al. Plant Dis. 88:1341, 2004. (2) N. A. Tisserat et al. Phytopathology 84:478, 1994.

Aspergillus flavus is a filamentous fungus that produces mycotoxins in many food and feed crops, such as maize (Zea mays L.). Isolates were analyzed for toxin production by nucleic acid profiles in an attempt to differentiate aflatoxigenic from nonaflatoxigenic isolates. A total of 41 aflatoxigenic and 34 nonalfatoxigenic isolates were included in the study. The isolates were evaluated initially using DNA amplification fingerprinting (DAF) without clear resolution of the groups. A weak association of aflatoxigenic isolates was observed, as evidenced by their clustering in 18 of 81 trees recovered from maximum parsimony analysis of binary characters derived from arbitrary signatures from amplification profiles (ASAP) data; nonaflatoxigenic isolates exhibited a pattern of paraphyletic laddering. Up to five markers unambiguously supported the aflatoxigenic isolate grouping, but the presence of alternative conflicting topologies in equally parsimonious trees precluded the observation of meaningful statistical support. With additional markers for genome of A. flavus, this method could be used to resolve toxigenic from nontoxigenic strains. This additional work could resolve aflatoxigenic isolates of A. flavus present on maize plants using ASAP, which would reduce labor intense costs and potentially lead to faster determination of resistant cultivars in breeding efforts.

Baseline information on the diversity and population densities of fungi collected from soil debris and cotton (Gossypium hirsutum L.) roots was determined. Samples were collected from Tifton, GA, and Starkville, MS containing cotton field soil treated with the nematicides 1,3-dichloroproprene (fumigant) and aldicarb (granules). A total of 10,550 and 13,450 fungal isolates were collected from these two study sites, respectively. Of this total, 34 genera of plant pathogenic or saprophytic species were identified. Pathogenic root fungi included Fusarium spp. (40% of all isolations), Macrophomina, Pythium, Rhizoctonia, and Sclerotium. Fusarium and Rhizoctonia were the most common fungal species identified and included F. oxysporum, F. verticillioides and F. solani, the three Fusarium species pathogenic on cotton plants. Population densities of Fusarium were not significantly different among locations or tissue types sampled. Macrophomina was isolated at greater numbers near the end of the growing seasons. Anastomosis groups of R. solani isolated from roots and soil debris included AG-3, -4, -7, 2-2, and -13 and anastomosis groups of binucleate Rhizoctonia included CAG-2, -3, and -5. Occurrences and frequency of isolations among sampling dates were not consistent. Fluctuations in the frequency of isolation of Rhizoctonia did not correspond with changes in frequency of isolation of the biological control fungus, Trichoderma. When individual or pooled frequencies of the mycobiota were compared to nematicide treatments, no specific trends occurred between treatments, application methods or rates. Results from this study show that use of 1,3-D and aldicarb in cotton fields does not significantly impact plant pathogenic fungi or saprophytic fungal populations. Thus cotton producers need not adjust seedling disease control measures when these two nematicides are used.

A 2-year (1999–2000) study was conducted at Starkville and Stoneville, MS to determine if the occurrence of the mycoflora varied on Roundup® Ready (transgenic) compared to conventional soybean (Glycine max) cultivars. A total of 7,658 fungal isolates were identified from the pod and seed tissues of four cultivars compared at growth stages R6 and R8. Ninety-nine percent of all fungi isolated were mitosporic fungi and ascomycetes. In both years, total fungal isolates from the two locations were greater from the pod (65%) than from seed (33%) tissues. Isolation frequency from conventional cultivars was 54% compared to 46% for the transgenic cultivars. The most common fungi identified that are reported pathogens of soybean included Alternaria, Cercospora, Cladosporium, Diaporthe, Fusarium and Verticillium spp. When main effects and interactions were compared among the frequency data for the fungal genera, significant differences occurred, but consistent trends were not noted. Isolation frequencies of Diaporthe spp. during the R6 growth stage, were significantly greater on the conventional than on the transgenic cultivars in both years of the study, but only at Starkville. Isolation frequencies from samples taken during the R8 growth stage were similar at both locations in 1999 and 2000.Fusarium spp. isolated at R6 and R8 growth stages from pod and seed tissues were significantly greater on conventional than on transgenic cultivars in 2000. Even though frequencies were often significantly different between the transgenic and conventional cultivars, the data was not consistent between locations, pod and seed tissues, or growth stages. The pod and seed mycoflora of transgenic and conventional soybean cultivars was, therefore, similar in Mississippi.

A review of the fungi associated with soybean seeds, pods, or flowers was conducted in North America. Species of Deuteromycetes are the most common fungi in each of the soybean flower organs followed by the Ascomycetes and Phycomycetes which comprise about one-fourth of the total mycoflora. Eighty genera and about 135 or more species occur in seeds, pods, or flowers. With regard to numbers of taxa from separate mycofloras, 63 genera and about 108 or more species occur in seeds, 65 genera and about 88 or more species occur in pods, and 36 genera and approximately 47 or more species occur in flowers. Most of the fungi which occur in flowers can be cultured from pods, and the majority of those fungi occur in seeds. Methods for and a key are provided for the identification of the 30 most important selected fungi.

In a field study evaluating the diversity and density of the soilborne mycobiota in a cotton (Gossypium hirsutum L.) production system, Macrophomina phaseolina (Tassi) Goidanich was isolated on potato dextrose agar from dark brown to black lesions on feeder and secondary roots. Multiple proliferations of feeder and secondary roots were also observed. Isolate RB 656 obtained from these lesions was tested for pathogenicity in the greenhouse by mixing 25 ml of 2-week-old cornmeal sand inoculum (3 g of cornmeal, 100 g of sand, and 20 ml of distilled water) with 5 liters of autoclaved soil (Leefield loamy sand, pH 6.2) per pot (40 × 200 cm). Control pots containing autoclaved soil alone were included for comparison. On 17 September, 10 cotton seeds of DPL 90 were sown per pot. Each treatment had five replications. Forty days after planting, plant heights from pots containing M. phaseolina were lower (14.8 cm) than those in the control pots (19.6 cm), but stand counts were similiar. On this date, four plants were left in each pot to allow the remaining seedlings to reach full maturity. On 9 December, the four plants per replicate pot were removed and roots evaluated for damage. Lesions similiar to those seen originally were observed on the secondary and feeder roots of the infested pots, and the characteristic proliferation of feeder and secondary roots was noted. Tap roots in pots containing M. phaseolina were smaller (11.6 cm) than in the control pots (18.6 cm). Isolate RB 656 was reisolated from the damaged roots in the treated pots. This is the first report of M. phaseolina on cotton in Georgia.