Fungi isolated from snap bean roots and rhizosphere soil where fungicides are not used included Fusarium oxysporum, Fusarium equiseti, Fusarium subglutinans, Fusarium camptoceras, Fusarium chlamydosporum, Fusarium verticillioides, Fusarium proliferatum, Fusarium acuminatum, Fusarium solani, Peyronellaea pinodella, Macrophomina phaseolina, and Glomerella guttata. Only P. pinodella, M. phaseolina, and F. oxysporum were isolated on symptomatic plants. These soilborne fungi are common pathogens of diverse host plants. Pathogenicity tests under controlled environment demonstrated that these fungi were pathogenic on snap beans. Subsequently, bacterial endophytes isolated from snap bean roots, papaya roots and stems, and dogwood stems were evaluated as potential biological control agents against these diverse fungi. All bacteria isolated, including Bacillus vallismortis (PS), Bacillus amyloliquefaciens (Psl), Bacillus subtilis (Prt), Bacillus thuringiensis (Y and IMC8), Enterobacter sp. (E), Stenotrophomonas sp. (B17A), and Serratia sp. (B17B) suppressed growth of the fungal pathogens in vitro and formed clear inhibition zones in petri dish dual cultures. Growth media taken from the inhibition zones suppressed growth of the fungal pathogens in the absence of the bacterial cells, suggesting that the bacteria released unidentified antagonistic biochemical substances into the media. This study constitutes an initial screening of endophytes as biological control agents against diverse fungal pathogens and forms a basis for the discovery of novel strains that can be further developed and integrated into disease management systems for diverse fungal pathogens. Isolates B. vallismortis (PS), B. amyloliquefaciens (Psl), B. subtilis (Prt), and B. thuringiensis (Y IMC8) exhibited the best performance as potential biological control agents paving the way for larger-scale in vivo studies and characterization of their interactions with fungal pathogens.
Jacqueline Joshua and Margaret T. Mmbaga
Margaret T. Mmbaga, and Roger J. Sauvé,
Twenty-five dogwood accessions (one Cornus kousa, three C. kousa × C. florida hybrids, and 21 C. florida) were characterized using amplified fragment length polymorphism. Among the C. florida accessions, four were named cultivars and 17 were selections from Tennessee State University's dogwood breeding program. Amplified fragment length polymorphism band profiles obtained from 13 EcoRI/MseI (+3/+3) primer pairs showed the presence of high genetic diversity between species and within the C. florida accessions. Each accession was distinctly different from each other, and the resistant clones clustered into distinct groups.
Suping Zhou, Roger J. Sauvé, and Margaret T. Mmbaga
A cold acclimatization mechanism regulated by the accumulation of mRNAs and proteins has been tentatively identified in japanese spurge (Pachysandra terminalis Sieb. & Zucc.). Two polypeptides and several cDNA fragments were observed in leaf tissue after acclimation. When these proteins were probed with type III fish antifreeze antibodies, an immune-cross reaction occurred. Nonacclimatized young leaves and stems of japanese spurge survived 20-minute exposures at -5 °C. Although newly emerged leaves and stems were damaged, plants resumed growth at higher temperatures. After acclimation by gradual cold treatments (4 to -5 °C), new proteins began to accumulate in young leaves and plants were more tolerant to extended treatments at -5 °C. Changes in accumulation of proteins and mRNA in leaf tissue of japanese spurge appear to be an adaptation mechanism to subfreezing conditions. This is the first report of the immune-cross reaction between antibodies of type III fish antifreeze proteins and plant proteins
Suping Zhou, Roger J. Sauvé, Margaret T. Mmbaga, and Chaim Frenkel
Leucanthemum maximum `Silver Princess' plants, that were gradually acclimated for 7 days at 10 °C followed by 28 days at 7 °C, were subjected to the following cold treatments: 30 days at 4 °C; 4 or 5 days at 0 °C and for 3 hours at –1 °C to identify cold inducible proteins that may be responsible for cold tolerance in this cold tolerant species. Change in antioxidant enzymes activity in fully expanded leaves was assessed after each treatment. Catalase activity began to increase after 30 days at 4 °C and reached its peak after a 5-day exposure to 0 °C. The activity of cellular glutathione peroxidase and glutathione reductase significantly increased after a 4-day exposure to 0 °C. Changes in activity of four active superoxide dismutase isoforms, one basic guaiacol peroxidase and two o-dianisine peroxidase isoforms were also detected following the full series of cold treatments (30 days at 4 °C; 4 or 5 days at 0 °C and for 3 hours at –1 °C).
Margaret T. Mmbaga, Lucas M. Mackasmiel, and Frank A. Mrema
Six biological control agents (BCAs) (two bacteria, two fungi, and two yeasts) that were previously shown to be effective against powdery mildew (Erysiphe pulchra) were tested for efficacy against Macrophomina phaseolina root rot on flowering dogwood (Cornus florida) in the greenhouse. Two of the bacterial isolates, Stenotrophomonas sp. (B17A) and Serratia sp. (B17B), were effective in controlling both macrophomina root rot and powdery mildew, similar to fungicide control thiophanate methyl, when roots were drenched with the six BCAs individually. In addition, the two bacterial BCAs improved plant growth with respect to stem diameter, stem length, dry weight, and green foliage compared with fungicide-treated plants or nontreated controls grown in sterile soil. These results confirm previous results in which B17A and B17B suppressed powdery mildew and also promoted plant growth in flowering dogwood. Although macrophomina root rot has been previously reported as a potential problem in flowering dogwood, especially in field conditions, simultaneous infection with macrophomina root rot and powdery mildew has not been previously reported. This study confirmed that M. phaseolina infection was characterized by stubby roots and black root lesions, and plants infected with both powdery mildew and macrophomina root rot had smaller root mass compared with fungicide-treated plants. Neither of the two pathogens killed their host plants, but compounded infections significantly reduced the plant root system and plant growth. The efficacy of the two bacterial isolates in controlling both powdery mildew and macrophomina root rot suggests their potential utilization in controlling both diseases in dogwood nursery production and in other plants that are hosts to both powdery mildew and macrophomina root rot. Plant growth promoted by the two BCAs may be attributed to powdery mildew and macrophomina root rot control, but comparisons between fungicide-treated plants and control plants not inoculated with BCAs or root rot pathogen suggested that the two BCAs may play a role as bio-stimulants in growth enhancement. These results also suggest that the two biocontrol agents are not phytotoxic to dogwood.
Margaret T. Mmbaga, Lucas A. Mackasmiel, and Frank A. Mrema
Macrophomina phaseolina was isolated from the crown region and roots of mature flowering dogwood (Cornus florida L.) trees in the landscape and nursery plantings. Although this pathogen has been reported in Cornus species, its occurrence and impact on C. florida has not been reported. Pathogenicity tests were conducted on dogwood seedlings, and all inoculated seedlings developed root necrotic lesions and no small lateral roots, whereas the non-inoculated control seedlings remained disease-free and developed numerous small roots. Seedlings inoculated with M. phaseolina exhibited numerous microsclerotia, but non-inoculated seedlings did not. In greenhouse experiments, plants inoculated on the stems near the soil line developed brown canker-like lesions and swellings around the inoculated area. These were not observed on non-inoculated plants.