Lactobacillus parafarraginis ZH1 producing anti‐yeast substances to improve the aerobic stability of silage

Enhancing the aerobic stability of silage is very important in silage production. The objective of this study was to compare the roles of a new bacterial strain Lactobacillus parafarraginis ZH1 with Lactobacillus plantarum Chikuso‐1 (LP) and Lactobacillus buchneri NCIMB 40788 (LB) in improving the aerobic stability of oat silage and identify the anti‐yeast substances produced by them. After ensiled for 45 days, either inoculated silages or control silage were of pH values lower than 4.2. The control and LP inoculated silage had poorer aerobic stability than LB and ZH1 inoculated ones (p < 0.05). ZH1 inoculated silage produced more acetic acid, benzoic acid and hexadecanoic acid than LB inoculated one and had the best aerobic stability (p < 0.05). In MRS medium, strain ZH1 produced more hexadecanoic acid than LP and LB strains, more benzoic acid than strain LP (p < 0.05), and the equal amount of benzoic acid to LB (p > 0.05). In addition, benzoic acid and hexadecanoic acid had low minimal inhibitory concentrations to target yeasts in the pure culture. In conclusion, L. parafarraginis ZH1 had stronger anti‐yeast potential and more effectively improved the aerobic stability of silage than other strains used in this study.     

Light,soil, and seedling characteristics associated with varying levels of competition in a red pine plantation

Red pine (Pinus resinosa Ait.) seedlings growing under differing levels of competition were evaluated during the fifth growing season following planting, and placed into low, moderate, or high tree classes, as a function of levels of competing vegetation. Tree growth, moisture status, and nutrition were monitored over the growing season. Additionally, site characteristics such as soil temperature and moisture, inorganic nitrogen concentration, mineralized soil nitrogen, and light were measured. Red pine seedlings growing under low competition had an absolute volume growth increase of 795% over the seedlings growing under the heaviest competition. The associated relative volume growth increase during the fifth growing season was 44%. Discriminant analysis was used to describe three classes of trees representing low, moderate, and high levels of competition. Trees growing under low competition had longer and heavier needles, but generally lower nutrient concentrations. Pre-dawn plant moisture status did not vary among competition levels. Soil variables indicated that, in general, the trees growing under low competition occupied warmer, drier, and less fertile microsites, as inorganic soil N and mean monthly mineralized NO₃-N and NH₄>-N tended to be lower on these microsites. The suite of independent variables was effective in classifying the model data into three tree competition classes, with percent correct classifications of 92, 75, and 100 for low (tree class one), moderate (tree class two), and heavy (tree class three) competition, respectively.     

Temporal Dynamics of the Biocontrol Agent Pseudomonas fluorescens Strain A506 in Flowers in Inoculated Pear Trees

ABSTRACT The colonization of individual flowers in mature pear orchards by Pseudomonas fluorescens strain A506 applied at different times during bloom was measured to determine the receptivity of flowers to colonization and the extent of intra-tree movement over time. Strain A506 populations in flowers open at inoculation were initially about 10(4) cells per flower and increased to approximately 10(6) cells per flower in flowers that were inoculated within about 5 days of opening. However, eventual populations decreased with further increases in flower age at inoculation to as few as about 10(3) cells per flower when inoculated flowers were more than 10 days old. Populations of strain A506 on flowers that opened after inoculation was initially very low at the time of petal expansion (<100 cells per flower) but increased rapidly with time after flower opening. The maximum population of strain A506 that developed on such flowers decreased with increasing time between inoculation and petal expansion; 10(4) to 10(5) cells of strain A506 eventually colonized flowers that opened within 7 days of inoculation, whereas fewer than 100 cells colonized flowers that opened 24 days or more after inoculation. Large total bacterial populations on A506-treated trees were associated with significant reductions in populations of Erwinia amylovora and reduced incidence of fire blight and severity of fruit russet.     

Occurrence of indole-3-acetic Acid-producing bacteria on pear trees and their association with fruit russet

ABSTRACT A relatively high percentage of epiphytic bacteria on pear leaf and fruit surfaces had the ability to produce indole-3-acetic acid (IAA) in culture media supplemented with tryptophan. While over 50% of the strains produced at least small amounts of IAA in culture, about 25% of the strains exhibited high IAA production as evidenced by both colorimetric and high-performance liquid chromatography analysis of culture supernatants. A majority of the strains that produced high amounts of IAA were identified as Erwinia herbicola (Pantoea agglomerans), while some strains of Pseudomonas syringae, Pseudomonas viridiflava, Pseudomonas fluorescens, Pseudomonas putida, and Rahnella aquaticus that produced high amounts of IAA also were found on pear. Fruit russeting was significantly increased in 39 out of 46 trials over an 8-year period in which IAA-producing bacteria were applied to trees compared with control trees. A linear relationship was observed between fruit russet severity and the logarithm of the population size of different IAA-producing bacteria on trees in the 30 days after inoculation, when normalized for the amount of IAA produced by each strain in culture. On average, the severity of fruit russet was only about 77% that on control trees when trees were treated at the time of bloom with Pseudomonas fluorescens strain A506, which does not produce IAA. Both total bacterial populations on pear in the 30-day period following full bloom and fruit russet severity varied greatly from year to year and in different commercial orchards over a 10-year period. There was a strong linear correlation between the logarithm of total bacterial population sizes and fruit russet severity.     

Contribution of Fimbrial and Afimbrial Adhesins of Xylella fastidiosa to Attachment to Surfaces and Virulence to Grape

ABSTRACT The role of fimbrial and afimbrial adhesins of Xylella fastidiosa in biofilm formation was assessed by visualization of cell aggregates of mutant strains after incubation on glass surfaces. FimA- or FimF- fimbrial mutants adhered as solitary cells at a slightly lesser frequency to glass surfaces than the parental strain; however, cell aggregates were not formed, unlike the wild-type strain. Conversely, whereas the XadA- and HxfB- nonfimbrial mutants also exhibited a much lower frequency of adherence to glass surfaces than the wild-type strain, most of the cells retained on the surfaces were in cell aggregates of different sizes, much like that of the parental strain. Neither fimbrial or afimbrial mutants formed a mature biofilm on the sides of flasks of broth cultures, unlike the dense biofilm formed by the wild-type strain. Although FimA- and FimF- mutants did not form cell aggregates on glass surfaces when incubated as individual strains, aggregates of a FimA- or FimF- mutant were observed when co-incubated with either a XadA- mutant or HxfB- mutant, respectively. These results are consistent with a model in which the fimbrial adhesins FimA and FimF are involved preferentially in cell-to-cell aggregate formation whereas the afimbrial adhesions XadA and HxfB preferentially contribute to initial cell binding to surfaces, whereupon further cell aggregation can occur. In each of five separate experiments, FimA, FimF, XadA, and HxfB mutants of X. fastidiosa all were less virulent to grape than the corresponding wild-type strain. Fimbrial and afimbrial mutants might produce a reduced biofilm within vessels of grape and, hence, be deficient in various cell-density-dependent traits required for movement through the plant and, thus, virulence.     

Genetic engineering of bacteria from managed and natural habitats

The genetic modification of bacteria from natural and managed habitats will impact on the management of agricultural and environmental settings. Potential applications include crop production and protection, degradation or sequestration of environmental pollutants, extraction of metals from ores, industrial fermentations, and productions of enzymes, diagnostics, and chemicals. Applications of this technology will ultimately include the release of beneficial agents in the environment. If safely deployed, genetically modified bacteria should be able to provide significant benefits in the management of environmental systems and in the development of new environmental control processes.     

Pseudomonas syringae Responds to the Environment on Leaves by Cell Size Reduction

ABSTRACT The length and volume of cells of the plant-pathogenic bacterium Pseudomonas syringae strain B728a were measured in vitro and with time after inoculation on bean leaf surfaces to assess both the effect of nutrient availability on the cell size of P. syringae and, by inference, the variability in nutrient availability in the leaf surface habitat. Cells of P. syringae harboring a green fluorescent protein marker gene were visualized by epifluorescence microscopy after recovery from leaves or culture and their size was estimated by analysis of captured digital images. The average cell length of bacteria grown on leaves was significantly smaller than that of cultured cells, and approached that of cells starved in phosphate buffer for 24 h. The average length of cells originally grown on King's medium B decreased from approximately 2.5 to approximately 1.2 mum by 7 days after inoculation on plants. Some decrease in cell size occurred during growth of cells on leaves and continued for up to 13 days after cell multiplication ceased. Although cultured cells exhibited a normal size distribution, the size of cells recovered from bean plants at various times after inoculation was strongly right-hand skewed and was described by a log-normal distribution. The skewness of the size distribution tended to increase with time after inoculation. The reduced cell size of P. syringae B728a on plants was readily reversible when recovered cells were grown in culture. Direct in situ measurements of cell sizes on leaves confirmed that most cells of P. syringae respond to the leaf environment by reducing their size. The spatial heterogeneity of cell sizes observed on leaves suggest that nutrient availability is quite variable on the leaf surface environment.     

Site-Directed Disruption of the fimA and fimF Fimbrial Genes of Xylella fastidiosa

ABSTRACT Xylella fastidiosa causes Pierce's disease, a serious disease of grape, citrus variegated chlorosis, almond and oleander leaf scorches, and many other similar diseases. Although the complete genome sequences of several strains of this organism are now available, the function of most genes in this organism, especially those conferring virulence, is lacking. Attachment of X. fastidiosa to xylem vessels and insect vectors may be required for virulence and transmission; therefore, we disrupted fimA and fimF, genes encoding the major fimbrial protein FimA and a homolog of the fimbrial adhesin MrkD, to determine their role in the attachment process. Disruption of the fimA and fimF genes in Temecula1 and STL grape strains of X. fastidiosa was obtained by homologous recombination using plasmids pFAK and pFFK, respectively. These vectors contained a kanamycin resistance gene cloned into either the fimA or fimF genes of X. fastidiosa grape strains Temecula1 or STL. Efficiency of transformation was sufficiently high ( approximately 600 transformants per mug of pFFK DNA) to enable selection of rare recombination events. Polymerase chain reaction and Southern blot analyses of the mutants indicated that a double crossover event had occurred exclusively within the fimA and fimF genes, replacing the chromosomal gene with the disrupted gene and abolishing production of the corresponding proteins, FimA or FimF. Scanning electron microscopy revealed that fimbriae size and number, cell aggregation, and cell size were reduced for the FimA or FimF mutants of X. fastidiosa when compared with the parental strain. FimA or FimF mutants of X. fastidiosa remained pathogenic to grapevines, with bacterial populations slightly reduced compared with those of the wild-type X. fastidiosa cells. These mutants maintained their resistance to kanamycin in planta for at least 6 months in the greenhouse.