• Proceedings of the Microbiological Society of Korea Conference
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• 2002.10a
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• pp.100-105
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• 2002

The diversity of epiphytic bacterial communities on deciduous oak tree (Quercus dentate Thunb.) leaves was examined both in the natural forest area with a clean air and in the industrial estate to assess effects of acidic deposition to the phyllosphere using 16S rDNA sequence data. In addition, acid-tolerant epiphytic bacterial communities were compared. A total of 78 epiphytic and 444 acid-tolerant clones were obtained from clone libraries, resulting in 20 and 17 phylotypes by analysis of restriction fragment length polymorphism (RFLP) for PCR-amplified 16S rDNA products. A low bacterial diversity in both areas was found. As tree leaves grow older, bacterial diversities were slightly increased in the level of subphylum. The community structure of epiphytic bacteria in both areas in April consisted of only two subphyla, $\beta-and\;\gamma-Proteobacteria$. In August two additional subphyla in both areas were found, but the composition was a little different, Acidobacteria and Cytophaga-Flexibacter-Bacteroids (CFB) group in the industrial estate and a -Proteobacteria and CFB group in the natural area, respectively. Acidobacteria could be an indicator of epiphytic bacteria for acidic deposition on plant leaves, whereas a -Proteobacteria be one of epiphytic bacteria that naturally survive on leaves that are not affected by acidic deposition. The acid-tolerant bacterial communities in April were composed of two subphyla, $\gamma-Proteobacteria$ and Low G+C gram-positive bacteria in both areas, and in August a-Proteobacteria was added to the community just in the natural forest area. The direct influence of acidic deposition on the acid-tolerant bacterial phylogenetic composition could not be detected in higher taxonomic levels such as subphylum, but at narrower or finer levels it could be observed by a detection of Xanthomonadales group of $\gamma-Proteobacteria$ just in the industrial estate.

• Microbiology and Biotechnology Letters
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• v.47 no.2
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• pp.303-309
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• 2019

Earlier studies by our group revealed that gallic acid in phytochemicals stimulated biofilm production in epiphytes, while caffeic acid in phytochemicals inhibited biofilm production in non-epiphytes. It is well documented that antimicrobial secretion by some epiphytic bacteria inhibits non-epiphytic bacterial growth on leaf surfaces. These selection criteria help plants choose their microbial inhabitants. Calcium and iron in phytochemicals also stimulate biofilm formation and thus, may be selection criteria adopted by plants with respect to their native epiphytic population. Furthermore, the processing of leaves during phytochemical extraction impacts the composition of the extract, and therefore its ability to affect bacterial biofilm formation. Computation of the Hurst exponent using biofilm thickness data obtained from the Ellipsometry of Brewster Angle Microscopic (BAM) images is an efficient tool for understanding the impact of phytochemicals on epiphytic and non-epiphytic populations when compared to fluorescent microscopy, scanning electron microscopy, and staining techniques. To the best of our knowledge, this is the first report that uses the Hurst exponent to elucidate the mechanism involved in plant microbe interaction.

• Korean Journal of Microbiology
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• v.37 no.4
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• pp.265-272
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• 2001

The diversity of acid-tolerant epiphytic bacterial communities on deciduous oak tree (Quercus dentate Thunb.) leaves was examined both in the natural forest area with a clean air and in the industrial estate to assess effects of acidic depositions to the phyllosphere using 16S rDNA sequence data. A total of 444 acid-tolerant epiphytic bacterial clones were obtained, resulting in 17 phylotypes by performing a analysis of restriction fragment length polymorphism (RFLP) for PCR-amplified 16S rDNA products. A very low diversity of dominating acid tolerant bacterial communities in both areas was found, just 2 subphyla groups, $\gamma$-Proteobacteria and low-G+C gram-positive bacteria. As tree leaves grow older, diversities of acid-tolerant bacteria on them significantly increased. The community structure of acid-tolerant epiphytic bacteria consisted of Pseudomonas and Enterobacteriaceae groups in the $\gamma$-Proteobacteria subphylum, and Streptococcaceae and Staphylococcus groups in the low-G+C gram-positive bacteria subphylum. The direct influence of acidic depositions on bacterial phylogenetic composition could not be detected especially when higher taxonomic levels such as subphylum, but at narrower or finer levels it could be observed by a detection of Xanthomonadales group belonged to the $\gamma$-Proteobacteria only in the industrial area and of Acetobacteraceae group belonged to the $\alpha$-Proteobacteria. There remains that these specific acid-tolerant epiphytic bacterial groups could be used as indicators for assessing effects of acidic depositions on the phyllosphere.

• Korean Journal of Microbiology
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• v.33 no.4
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• pp.262-266
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• 1997

Total, direct viable count, and acid-tolerant epiphytic bacterial population sizes were quantified on leaves of chestnut tree (Castanea crenata S. et Z.) near Taejon Industrial Estate affected by acid precipitation and deposition as well as in the clean natural forest area, Mt. Kyejok, in Taejon city from August 1996 to August 1997. Geometric mean numbers of total, direct viable count, and acid-tolerant epiphytic bacteria were $9.9{\times}10^5cell/cm^2$, $1.6{\times}10^6cell/cm^2$, and $7.1{\times}10^3cfu/cm^2$ respectively, being 1.5, 2, and 2.6 times those in the clean area. Acid-tolerant epiphytic bacterial numbers at pH 5.6 by MPN method were $3.3{\times}10^4$ in the industrial area, about the same as the number, $3.4{\times}10^4MPN/cm^2$, of the clean area. Acid-tolerant bacterial number at pH 4.0 was $1.9{\times}10^{-1}MPN/cm^2$ in the industrial area, whereas none was detected in the clean area. Acid-tolerant bacteria at pH 3.0 were not detected at all in the industrial area as well as in the clean area. Epiphytic bacterial population sizes were generally the greatest in May when leaves are emerged and grew hut the lowest in November when defoliation occurs. These results showed that air pollutant deposition on leaves did not cause a decrease of epiphytic bacteria at least and acid deposition on leaves did cause an increase of acid-tolerant bacteria.

• Korean Journal of Microbiology
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• v.37 no.1
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• pp.61-65
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• 2001

Epifluorescence microscopy was used to observe epiphytic bacteria directly on plant leaf surfaces as well as indirectly in the leaf liberating solution by staining with fluorochromes of 4',6-diamidino-2-phenylindole (DAPI) and acridine orange(AO). Epiphytic bacteria could not be well observed on the leaf surface by staining with AO due to an intrusive orange or red background fluorescence. However, DAPI gave us clear epifluorescent images of the bacteria on the leaf. On the contrary, epiphytic bacteria in the liberating leaf solution were well observed on filters stained by both types of fluorochrome, although DAPI showed better fluorescent images than AO and not necessarily required a washing step of the filters stained. The optimum conditions of the DAPI stains were 5 $\mu$g/ml for 5 min both for leaves and for filters of the liberating solution. It was confirmed that a critical step in the epifluorescence microscopy of leaf surfaces was to minimize release of water from the leaf. For this, the stained leaf samples were put on a filter paper, kept in a dry oven at $70^{\circ}C$ for 2 min instead of air-drying, and then immediately observed by epifluorescence microscopy. The established technique was applied to enumerate epiphytic bacteria on oak tree leaf surfaces.

• Asian-Australasian Journal of Animal Sciences
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• v.18 no.6
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• pp.807-811
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• 2005

This experiment examined the characteristics of fermented juice of epiphytic lactic acid bacteria (FJLB) prepared by the addition of glucose, sucrose and molasses as a fermentation substrate. The effect of FJLB on the fermentative quality and changes in chemical composition during fermentation of guineagrass silage were also investigated. The pH value of the silages treated with FJLB rapidly decreased, and reached to the lowest value within 7 days of start of fermentation, as compared to the control. The number of lactic acid bacteria (LAB) in the treated silages increased for the first 3 days, thereafter the number of LAB declined gradually up to the end of the experiment. Silages treated with FJLB had larger populations of LAB than the control. Ammonia-nitrogen production increased throughout the ensiling period, which in the control and no-sugar added FJLB silages were higher than the other treated silages. Lactic acid levels varied with the time of ensiling and among the silage treatments. For any sugar FJLB treated silages, the lactic acid increased initially, and then slightly reduced to less than 50 g/kg of dry matter until 49 days after ensiling, except the silage treated with glucose added FJLB. Nevertheless, lactic acid content of the control decreased constantly from the beginning of ensiling and was not found after 35 days. Moreover, acetic acid content increased throughout the ensiling period. All the FJLB treated silages had significantly (p<0.05) lower pH and ammonia-nitrogen content, while significantly (p<0.05) higher lactic acid content and V-score value compared with the control. This study confirmed that the applying of FJLB with any sugar substrate improved fermentative quality of silage.

• Proceedings of the Korean Society of Plant Pathology Conference
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• 1999.04a
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• pp.43.4-44
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• 1999

• Animal Bioscience
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• v.35 no.12
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• pp.1860-1870
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• 2022

Objective: The observation that temperate C₃ and tropical C₄ forage silages easily produce large amounts of ethanol or acetic acid has puzzled researchers for many years. Hence, this study aimed to assess the effects of epiphytic microbiota from C₃ forages (Italian ryegrass and oat) on fermentative products and bacterial community structure in C₄ forage (sorghum) silage. Methods: Through microbiota transplantation and γ-ray irradiation sterilization, the irradiated sorghum was treated: i) sterile distilled water (STSG); ii) epiphytic microbiota from sorghum (SGSG); iii) epiphytic microbiota from Italian ryegrass (SGIR); iv) epiphytic microbiota from oat (SGOT). Results: After 60 days, all the treated groups had high lactic acid (>63.0 g/kg dry matter [DM]) contents and low pH values (<3.70), acetic acid (<14.0 g/kg DM) and ammonia nitrogen (<80.0 g/kg total nitrogen) contents. Notably, SGIR (59.8 g/kg DM) and SGOT (77.6 g/kg DM) had significantly (p<0.05) higher ethanol concentrations than SGSG (14.2 g/kg DM) on day 60. After 60 days, Lactobacillus were predominant genus in three treated groups. Higher proportions of Chishuiella (12.9%) and Chryseobacterium (7.33%) were first found in silages. The ethanol contents had a positive correlation (p<0.05) with the abundances of Chishuiella, Acinetobacter, Stenotrophomonas, Chryseobacterium, and Sphingobacterium. Conclusion: The epiphytic bacteria on raw materials played important roles in influencing the silage fermentation products between temperate C₃ and tropical C₄ forages. The quantity and activity of hetero-fermentative Lactobacillus, Chishuiella, Acinetobacter, Stenotrophomonas, Chryseobacterium, and Sphingobacterium may be the key factors for the higher ethanol contents and DM loss in silages.

• Plant Disease and Agriculture
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• v.5 no.1
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• pp.27-33
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• 1999

Frost injury of crops is closely related to the epiphytic population dynamics of ice nucleation-active (INA) bacteria, and the injury can be reduced by decreasing the INA bacterial population. In order to predict the epiphytic population of INA bacteria on crops, a rapid and accurate detection method has to be developed. In the previous report, we produced some antibodies against INA proteins purified from the outer membrane of INA bacteria. However it was difficult to produce the antibodies because the purification procedures of the INA proteins were complicated, and the final yield was too low. We designed a specific peptide from the N-terminal region of INA protein by computer analysis and synthesized the peptide in vitro in this experiment. The peptide sequence was Asp-Ser-Por-Leu-Ser-Leu-His-Ala-Asp, that is corresponding to the highly conserved region in several INA proteins, with predicted beta turn, coiling, and hydrophilic region. A polyclonal anti-INA peptide antiserum produced specifically recognized INA bacteria as few as 10 colony-forming units (CFU) in the ELISA reactions and did not respond to other non-INA bacteria. Serological specificity of the anti-INA peptide antiserum will facilitate the forecasting of the INA bacterial population dynamics on crops.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.6
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• pp.920-924
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• 2007

Two experiments were carried out on a laboratory scale. The first involved a study of the effect of green tea on characteristics of fermented juice of epiphytic lactic acid bacteria (FJLB). FJLB was treated with 50 g/L of green tea products as follows: new shoot powder (FJLB+N), leaf powder (FJLB+L), commercial powder (FJLB+P), sterilized new shoot powder (FJLB+SN), sterilized leaf powder (FJLB+SL) or sterilized commercial powder (FJLB+SP). FJLB without any additive was also prepared (Untreated FJLB). After incubation, the number of microorganisms in FJLB were studied. Subsequently, these FJLB were applied at 10 ml/kg to chopped rhodesgrass to study their effects on fermentation. Compared with untreated FJLB, the addition of green tea increased (p<0.05) lactic acid bacteria (LAB) and also aerobic bacteria counts in FJLB. At 60 d of ensiling, all the FJLB treated silages were well preserved, pH and butyric acid content were lower (p<0.001) and lactic acid was higher (p<0.001) than that of the control. Lactic acid content was significantly higher (p<0.001) with treated FJLB than with untreated FJLB. FJLB treated with sterilized green tea decreased (p<0.001) the pH and the lactic acid content was higher (p<0.001) than that in the unsterilized green tea silages.