Agricultural antibiotics are widely used to inhibit the growth of phytopathogenic bacteria involved in plant diseases. However, continuous antibiotic overuse in crop production may lead to the development of antibiotic resistance in phytopathogenic bacteria. This study was conducted to evaluate the resistance to three different agricultural antibiotics (oxytetracycline+streptomycin, streptomycin, and validamycin A) in 91 strains of phytopathogenic bacteria including Pectobacterium carotovorum, Pseudomonas syringae pv. actinidiae, Clavibacter michiganensis subsp. michiganensis, C. michiganensis subsp. capsici, and Xanthomonas arboricola pv. pruni. Bacterial growth in the presence of various concentrations of validamycin A was also assessed spectrophotometrically by analyzing the optical density. All strains did not grow when the cells were exposed to oxytetracycline+streptomycin or 100× of streptomycin. However, among the 91 strains, 4% and 2% strains showed bacterial growth at the concentrations of 1× and 10× of streptomycin, respectively. Furthermore, 97%, 93%, and 73% strains were resistant to the 1×, 10×, and 100× of validamycin A, respectively, and especially, P. carotovorum contained the highest resistance to the validamycin A. Minimum bactericidal concentration values of validamycin A did not correlate with the patterns of agricultural antibiotic resistance. Further studies are needed to understand the incidence and development of antibiotic resistance in phytopathogenic bacteria.
As a member of ectomycorrhizal fungi, Tricholoma matsutake has a symbiotic relationship with its host, Pinus densiflora. To cultivate T. matsutake artificially, the co-cultivation of T. matsutake mycelia and bacteria from shiro was introduced. In this study, bacteria were isolated from soil samples in Bonghwa-gun, and seven bacterial isolates (B22_7_B05, B22_7_B06, B22_7_B07, B22_7_B08, B22_7_B10, B22_7_B13, and B22_7_B14) promoted the growth of T. matsutake mycelia (147.48, 232.11, 266.72, 211.43, 175.17, 154.62, and 177.92%, respectively). Sequencing of the 16S rRNA region of the isolated bacteria was performed. B22_7_B05 and B22_7_B10 were identified as Bacillus toyonensis, B22_7_B06 and B22_7_B08 as Paenibacillus taichungensis, B22_7_B07 and B22_7_B14 as P. gorilla, and B22_7_B13 as P. odorifer. These bacterial isolates were associated with the shiro community and are expected to contribute to the cultivation of T. matsutake.
Antagonistic microorganisms against $Rhizoctonia$$solani$ were isolated and their antifungal activities were investigated. Two hundred sixteen bacterial isolates were isolated from various soil samples and 19 isolates were found to antagonize the selected plant pathogenic fungi with varying degrees. Among them, isolate C9 was selected as an antagonistic microorganism with potential for use in further studies. Treatment with the selected isolate C9 resulted in significantly reduced incidence of stem-segment colonization by $R.$$solani$ AG2-2(IV) in Zoysia grass and enhanced growth of grass. Through its biochemical, physiological, and 16S rDNA characteristics, the selected bacterium was identified as $Bacillus$$subtilis$ subsp. $subtilis$. Mannitol (1%) and soytone (1%) were found to be the best carbon and nitrogen sources, respectively, for use in antibiotic production. An antibiotic compound, designated as DG4, was separated and purified from ethyl acetate extract of the culture broth of isolate C9. On the basis of spectral data, including proton nuclear magneric resonance ($^1H$ NMR), carbon nuclear magneric resonance ($^{13}C$ NMR), and mass analyses, its chemical structure was established as a stereoisomer of acetylbutanediol. Application of the ethyl acetate extract of isolate C9 to several plant pathogens resulted in dose-dependent inhibition. Treatment with the purified compound (an isomer of acetylbuanediol) resulted in significantly inhibited growth of tested pathogens. The cell free culture supernatant of isolate C9 showed a chitinase effect on chitin medium. Results from the present study demonstrated the significant potential of the purified compound from isolate C9 for use as a biocontrol agent as well as a plant growth promoter with the ability to trigger induced systemic resistance of plants.
To investigate the effects of immunization against somatostatin (SRIF) on growth rate, feed efficiency and carcass quality; forth-eight Yorkshire gilts ($age=37.5{\pm}4.3d,\;wt=8.2{\pm}1.6kg$) were randomly assigned to one of the following three treatments (1) control, (2) bovine serum albumin (BSA) and (3) SRIF. Cyclic SRIF was conjugated to BSA as the antigen containing 1 mg of SRIF diluted in 3 ml of saline. The conjugate was injected subsutaneously together with bacterial cell protein (BP) adjuvant on both sides of the neck of each gilt as the initial injection with three subsequent booster injections. Throughout the experiment all pigs were fed ad libitum a corn-soy diet containing 20% protein. Body weight and feed intake were measured on a weekly basis. All pigs in the experiment were slaughtered when they approached 101 kg body weight on the weekly weigh day. After slaughter, carcass parameters were analyzed to assess carcass quality. Results revealed that there were no differences among SRIF, BSA and control treatments for average daily gain, feed efficiency and feed intake during the first 5 wk of the experiment and from 6 wk to slaughter. The results for carcass analysis indicated that active immunization against SRIF had no effect on fat content, lean yield, water content and Canadian carcass index These data, collectively, suggest that the protocol employed in the present investigation for active immunization against SRIF is not an effective method for the enhancement of pig growth and improvement of feed efficiency and carcass quality.
This study was conducted to compare the effects of dietary supplementation of Sodium Gluconate (SG), Mannan Oligosaccharide (MOS) and Potassium Diformate (PDF) on growth performance and small intestinal morphology in nursery piglets. One hundred forty four female piglets ($11.69{\pm}0.71\;kg$) were divided into 4 treatments with six replicates of six pigs each. The pigs received a control diet or diets supplemented with SG, MOS and PDF at 2,500, 3,000 and 8,000 ppm; respectively, for 6 weeks. Supplementation of SG, MOS or PDF increased final body weight, average daily gain and tended to improve feed to gain ratio (p = 0.02, 0.04 and 0.16; respectively), other than average daily feed intake, intestinal pH and the bacterial populations were not influenced by the dietary treatments. SG significantly decreased the ammonia concentration in the caecum (p<0.05) and supplementation of SG, MOS or PDF tended to increase lactic acid and total short chain fatty acid concentration in the caecum (p = 0.08, 0.09; respectively), in addition SG, MOS or PDF slightly increased butyric acid concentration in the caecum (p = 0.14). SG highly significant increased the villous height in jejunum (p<0.01) and supplementing SG, MOS or PDF significantly increased crypt depth in jejunum (p<0.05), moreover, PDF significantly increased villous height and crypt depth ratio in jejunum (p<0.05) compared with control. The dietary treatments did not influence villous height and crypt depth in duodenum and villous height in jejunum (p>0.05). It can be concluded that supplementing SG, MOS or PDF as a feed additive has the potential to improve the growth performance, the intestinal lactic acid bacteria population, intestinal short-chain fatty acid concentration and the intestinal morphology of pigs.
The purpose of this study is to investigate the antimicrobial effect of mugwort (Artemisia asiatica Nakai) on the rice cake and bread preservation, and to identify their antimicrobial compounds. The mugwort extracts showed complete inhibition on the growth of Bacillus subtilis and Staphylococcus aureus at 250 $\mu\textrm{g}$/ml level. Antimicrobial activi쇼 of mugwort extract were stronger than that of commercial antimicrobial agent. Five % of sodium propionate solution showed complete inhibition on the growth of B. subtilis, E. coli and S. aureus, but L. plantarum was inhibited 50.87% at the same concentration. When various amounts of freeze-dried mugwort powder were added in sulgis (steamed rice cake), 3% ssooksulgi (mugwort powder added sulgi) had quite lower level of total bacterial count (5.5$\times$$10^/5 CFU/g) compared with the control group (1.4$\times$$10^/7 CFU/g) at ambient temp. (30$\pm$1$^{\circ}C$) after 72 hr. Three % addition of mugwort showed 2 days extention of shelf-life of rice cake. The sensory qualities of ssooksulgi has no significant difference in moistness, consistency, cohesiveness, afterswallowing and overall quality compared with control group. Ssooksulgi with 3% of mugwort powder had the best overall quality in sensory test. The methanol extract of 500 $\mu\textrm{g}$/ml of mugwort could lead the successful retardation of the growth of putrefactive microorganism during the incubation of rice cake at 37$^{\circ}C$ for 24 hr. On the other hand, coumarin (Sigma) had 54% inhibitory effect at 500 $\mu\textrm{g}$/ml level, and (E,E)-2,4-decadienal completely inhibited the growth of putrefactive microorganism of whitesulgi at 100 $\mu\textrm{g}$/ml level during the incubation at 37$^{\circ}C$ for 48 hr.48 hr.
Huh, Yoon Sun;Lee, Joung Kwan;Kim, Ik Jei;Kang, Bo Goo;Lee, Ki Yeol
Journal of Plant Biotechnology
/
v.42
no.2
/
pp.111-116
/
2015
Interest and great demand for blueberry (Vaccinium corymbosum) have increased, as V. corymbosum is now one of the most economically important crops in Korea. It is expected that blueberry production and the area planted for cultivation will increase consistently in the years ahead because of high profitability and the consumer's demand for healthy ingredients. Effective mass production of blueberry is urgently needed for commercial cultivation establishment, but a main limitation is lack of a propagation system that produces a disease-free plant material for commercial plantation. A large amount of research has focused entirely on developing tissue culture techniques for blueberry propagation. However, controlling fungal and bacterial contamination of woody plant material is extremely difficult. Our study was conducted to investigate the effect of biocide addition during the in vitro culture of blueberry on plantlet growth and contamination occurrence. Four biocides, including Plant Preservative Mixture ($PPM^{TM}$), vancomycin, nystatin and penicillin G, were used in varying concentrations during the in vitro propagation of blueberry. When nystatin was added into the medium at low concentrations, the overall growth of blueberry plantlets was retarded. Addition of vancomycin and penicillin G in high concentrations decreased contamination but induced plantlet mortality. On the other hand, when 1ml/L $PPM^{TM}$ was added, the growth characteristics of blueberry plantlets did not significantly differ from non-treatment (control), and the contamination occurrence rate was very low. From these results, we found that the addition of the appropriate biocide could provide an effective method to reduce contamination in the culture process, thereby raising in vitro production efficiency.
Anaerobic ammonium oxidation (AMX) is a cost-efficient biological nitrogen removal process. The coexistence of ammonia-oxidizing bacteria (AOB) in an AMX reactor is an interesting research topic as a nitrogen-related bacterial consortium. In this study, a sequencing batch reactor for AMX (AMX-SBR) was operated with a conventional activated sludge. The AOB in an AMX bioreactor were identified and quantified using terminal restriction fragment length polymorphism (T-RFLP) and real-time qPCR. A T-RFLP assay based on the ammonia monooxygenase subunit A (amoA) gene sequences showed the presence of Nitrosomonas europaea-like AOB in the AMX-SBR. A phylogenetic tree based on the sequenced amoA gene showed that AOB were affiliated with the Nitrosomonas europaea/mobilis cluster. Throughout the enrichment period, the AOB population was stable with predominant Nitrosomonas europaea-like AOB. Two OTUs of amoA_SBR_JJY_20 (FJ577843) and amoA_SBR_JJY_9 (FJ577849) are similar to the clones from AMX-related environments. Real-time qPCR was used to quantify AOB populations over time. Interestingly, the exponential growth of AOB populations was observed during the substrate inhibition of the AMX bacteria. The specific growth rate of AOB under anaerobic conditions was only 0.111 d-1. The growth property of Nitrosomonas europaea-like AOB may provide fundamental information about the metabolic relationship between the AMX bacteria and AOB.
The growth-inhibitory activity of Cassia tora seed-derived materials against seven intestinal bacteria was examined in vitro, and compared with that of anthraquinone, anthraflavine, anthrarufin, and 1-hydroxyanthraquinone. The active constituent of C. tore seeds was characterized as quinizarin, using various spectroscopic analyses. The growth responses varied depending on the compound, dose, and bacterial strain tested. At 1 mg/disk, quinizarin exhibited a strong inhibition of Clostridium perfringens and moderate inhibition of Staphylococcus aureus without any adverse effects on the growth of Bifidobacterium adolescentis, B. bifidum, B. longum, and Lactobacillus casei. Furthermore, the isolate at 0.1 mg/disk showed moderate and no activity against C. perfringens and S. aureus. The structure-activity relationship revealed that anthrarufin, anthraflavine, and quinizarin moderately inhibited the growth of S. aureus. However. anthraquinone and 1-hydroxyanthraquinone did not inhibit the human intestinal bacteria tested. As for the morphological effect of 1 mg/disk quinizarin, most strains of C. perfringens were damaged and disappeared, indicating that the strong activity of quinizarin was morphologically exhibited against C. perfringens. The inhibitory effect on aflatoxin $B_1$ biotransformation by anthraquinones revealed that anthrarufin ($IC_50,\;11.49\mu\textrm{M}$) anthraflavine ($IC_50,\;26.94\mu\textrm{M}$), and quinizarin ($IC_50,\;4.12\mu\textrm{M}$), were potent inhibitors of aflatoxin ${B_1}-8,9-epoxide$ formation. However, anthraquinone and 1-hydroxyanthraquinone did not inhibit the mouse liver microsomal sample to convert aflatoxin $B_1$ to aflatoxin ${B_1}-8,9-epoxide$. These results indicate that the two hydroxyl groups on A ring of anthraquinones may be essential for inhibiting the formation of aflatoxin ${B_1}-8,9-epoxide$. Accordingly, as naturally occurring inhibitory agents, the C. tora seed-derived materials described could be useful as a preventive agent against diseases caused by harmful intestinal bacteria, such as clostridia, and as an inhibitory agent for the mouse liver microsomal conversion of aflatoxin $B_1$ to aflatoxin ${B_1}-8,9-epoxide$.
Saad, N.;Urdaci, M.;Vignoles, C.;Chaignepain, S.;Tallon, R.;Schmitter, J.M.;Bressollier, P.
Journal of Microbiology and Biotechnology
/
v.19
no.12
/
pp.1635-1643
/
2009
The aim of this study was to provide new insight into the mechanism whereby the housekeeping enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) locates to cell walls of Lactobacillus plantarum 299v. After purification, cytosolic and cell wall GAPDH (cw-GAPDH) forms were characterized and shown to be identical homotetrameric active enzymes. GAPDH concentration on cell walls was growth-time dependent. Free GAPDH was not observed on the culture supernatant at any time during growth, and provoked cell lysis was not concomitant with any reassociation of GAPDH onto the cell surface. Hence, with the possibility of cw-GAPDH resulting from autolysis being unlikely, entrapment of intracellular GAPDH on the cell wall after a passive efflux through altered plasma membrane was investigated. Flow cytometry was used to assess L. plantarum 299v membrane permeabilization after labeling with propidium iodide (PI). By combining PI uptake and cw-GAPDH activity measurements, we demonstrate here that the increase in cw-GAPDH concentration from the early exponential phase to the late stationary phase is closely related to an increase in plasma membrane permeability during growth. Moreover, we observed that increases in both plasma membrane permeability and cw-GAPDH activity were delayed when glucose was added during L. plantarum 299v growth. Using a double labeling of L. plantarum 299v cells with anti-GAPDH antibodies and propidium iodide, we established unambiguously that cells with impaired membrane manifest five times more cw-GAPDH than unaltered cells. Our results show that plasma membrane permeability appears to be closely related to the efflux of GAPDH on the bacterial cell surface, offering new insight into the understanding of the cell wall location of this enzyme.
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