• Biotechnology and Bioprocess Engineering:BBE
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• v.10 no.4
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• pp.322-328
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• 2005

Acylhomoserine lactones (AHLs) are known to be the triggering molecules in the quorum sensing mechanism of many gram-negative bacteria. In order to detect AHL inhibitors that are potential biofilm inhibitors, a convenient and sensitive bioassay was developed based on the $\beta$-galactosidase activity ($\beta$-GAL) of a recombinant Agrobacterium tumefaciens strain. A series of commercially available AHLs were tested for inducing $\beta$-GAL at varying concentrations in agar-plate and liquid cultures of the reporter strain. All AHLs tested exhibited a concentration­dependent induction, and octanoyl homoserine lactone (OHL) showed the highest sensitivity with a detection limit of 0.1 nM in the liquid culture assay. When fimbrolide, a known quorum sensing inhibitor, was added, induction of $\beta$-GAL by OHL was repressed. The repression at a constant OHL concentration was dependent on the fimbrolide concentration with the detection limit below 1 ppm, indicating that this assay is a sensitive method for screening AHL inhibitors.

• Journal of Microbiology and Biotechnology
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• v.26 no.3
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• pp.521-529
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• 2016

Extensive use of antibiotics over recent decades has led to bacterial resistance against antibiotics, including gentamicin, one of the most effective aminoglycosides. The emergence of resistance is problematic for hospitals, since gentamicin is an important broad-spectrum antibiotic for the control of bacterial pathogens in the clinic. Previous study to identify gentamicin resistance genes from environmental samples have been conducted using culture-dependent screening methods. To overcome these limitations, we employed a metagenome-based culture-independent protocol to identify gentamicin resistance genes. Through functional screening of metagenome libraries derived from soil samples, a fosmid clone was selected as it conferred strong gentamicin resistance. To identify a specific functioning gene conferring gentamicin resistance from a selected fosmid clone (35-40 kb), a shot-gun library was constructed and four shot-gun clones (2-3 kb) were selected. Further characterization of these clones revealed that they contained sequences similar to that of the RNA ligase, T4 rnlA that is known as a toxin gene. The overexpression of the rnlA-like gene in Escherichia coli increased gentamicin resistance, indicating that this toxin gene modulates this trait. The results of our metagenome library analysis suggest that the rnlA-like gene may represent a new class of gentamicin resistance genes in pathogenic bacteria. In addition, we demonstrate that the soil metagenome can provide an important resource for the identification of antibiotic resistance genes, which are valuable molecular targets in efforts to overcome antibiotic resistance.

• Journal of Oral Medicine and Pain
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• v.24 no.2
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• pp.181-187
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• 1999

The most important progress in diagnostic sciences is the increased sensitivity and specificity in diagnostic procedures due to the development of newer micromethodologies and increasing availability of immunological and molecular biological reagents. The outcome of researches in this field has already provided DNA probes and antibodies which can be used for diagnosing various kinds of diseases including inherited ones. This development can be also applied to diagnose diseases in oral and maxillofacial regions. Technological advances have yielded highly sensitive test methodologies so that low analyte concentration and small sample volume are no longer limiting factors. Therefore, saliva can be useful test fluid for an array of analytes. Salivary constituents of diagnostic significance include steroid hormones, antibodies, drugs, and tumor markers. Of the proteins present in saliva, viral-specific immunoglobulins are of the greatest diagnostic interest. The development of conjugates and antigens by recombinant DNA technique and peptide synthesis is necessary for clinical application. Several kits developed for the purpose of blood testing should be modified to permit their application to saliva. The final practical outcome of researches in diagnostic sciences will be various diagnostic agents which can be used for detection of bacteria and viruses, screening and diagnosis of diseases, genetic screening for forensic individual identification. For these purposes, collaboration researches and development between institutions and companies are essential.

• Journal of Microbiology and Biotechnology
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• v.23 no.6
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• pp.850-855
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• 2013

Finding new antimicrobial activities by functional metagenomics has been shown to depend on the heterologous host used to express the foreign DNA. Therefore, efforts are devoted to developing new tools for constructing metagenomic libraries in shuttle vectors replicatable in phylogenetically distinct hosts. Here we evaluated the use of the Escherichia coli-Bacillus subtilis shuttle vector pHT01 to construct a forest-soil metagenomic library. This library was screened in both hosts for antimicrobial activities against four opportunistic bacteria: Proteus vulgaris, Bacillus cereus, Staphylococcus epidermidis, and Micrococcus luteus. A new antibacterial activity against B. cereus was found upon screening in B. subtilis. The new antimicrobial agent, sensitive to proteinase K, was not active when the corresponding DNA fragment was expressed in E. coli. Our results validate the use of pHT01 as a shuttle vector and B. subtilis as a host to isolate new activities by functional metagenomics.

• The Korean Journal of Mycology
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• v.21 no.2
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• pp.112-119
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• 1993

Various microorganisms secreting plant growth regulators were screened from the 100 microbial isolates including bacteria, actinomycetes and fungi. The isolates showing distict influence on the plant growth were identified as Aspergillus niger. The germinations of Raphanus and Cucubis seeds were completely inhibited by the culture filtrates of A. niger KK, A. niger KKS and A. niger ATCC 9462. The culture filtrates of the three strains also inhibited the formation and development of roots and hypocotyls of Raphanus. The culture filtrates of A. niger ATCC 26550 induced the hypocotyl curvature of Raphanus like plant hormone-auxin and at the same time caused the necrosis of the whole leaves.

• Korean Journal of Organic Agriculture
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• v.24 no.4
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• pp.945-955
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• 2016

To date, chemical managements of plant bacterial diseases are complicated by limitations on use of antibiotics in agriculture, antibiotic resistance development, and limited efficacy of alternative control agents. In this study, thus, we performed screening of eco-friendly farming material (notice no. 2-4-064) containing tannic acid as new antibacterial-activity against 7 plant bacterial pathogens: Pectobacterium carotovorum subsp. carotovorum (Pcc), Ralstonia solanacearum (Rs), Acidovorax avenae subsp. citrulli (Aac), Xanthomonas cirti pv. citri (Xcc), Erwinia pyrifoliae (Ep), Clavibacter michiganensis subsp. michiganensis (Cmm), and Streptomyces scabies (Sc), Initial screening of antibacterial effects of eco-friendly farming material was performed using the paper disk diffusion method and co-cultivation in broth culture. Tannic acid based eco-friendly farming material showed inhibitory effect against Pcc, Rs, Aac, Xcc, Cmm, and Ss, whereas it did not show inhibition zone against Ep. However, when it tested by co-cultivation in broth culture, it showed strong inhibition effect against all pathogens that declined growth curve compared to bacterial pathogen only. Interestingly, when we observed morphological changes on those pathogens by SEM, cell morphologies of 7 pathogens were slightly changed that seem to be malfunction in their cell envelope.

• Microbiology and Biotechnology Letters
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• v.35 no.4
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• pp.278-284
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• 2007

A novel screening strategy for salt-resistant antimicrobial peptides from a M13 peptide library was developed. Fusion of MSI-344, a magainin derivative and indolicidin to pIII coat proteins did not significantly affect viability of the recombinant phages, which indicated that the pIII could neutralize toxicity of the antimicrobial peptides and therefore it is possible to construct antimicrobial peptide library in Escherichia coli. On the basis of the conserved sequence of ${\alpha}$-helical antimicrobial peptides, a semi-combinatorial peptide library was constructed in which the peptides were displayed by pIII. To remove hemolytic activity from the library, the phages bound to red blood cells were removed, and the subtracted phage library was screened for binding to target bacteria Pseudomonas aeruginosa and Staphylococcus aureus under high salt concentrations. The screened peptides showed relatively low antimicrobial activity against the target bacteria. However, antimicrobial activities of the screened peptides P06 and S18 were not affected by the cation concentrations of 150 mM $Na^+$, 2 mM $Mg^{2+}$ and 2 mM $Ca^{2+}$ without significant hemolytic activity. This screening strategy that is based on binding capacity to target cells provides new potential to develop salt-tolerant antimicrobial peptides.

• Natural Product Sciences
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• v.4 no.1
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• pp.32-37
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• 1998

The water extracts of oriental herbal medicines which have been clinically used to treat bacterial infections in Korea were screened for in vitro antibacterial activity by the paper disc assay method. Two Gram positive bacteria, Staphylococcus aureus SG511, Bacillus subtilis ATCC 6633 and two Gram negative bacteria, Escherichia coli 055, Pseudomonas aeruginosa 9027 were used as test organisms. Among 83 of the extracts tested, 25 were active against Staphylococcus aureus SG511, 9 were active against Bacillus subtilis ATCC 6633, while none showed inhibitory activity against Eschelichia coli 055 and Pseudomonas aeruginosa 9027. Among them, Hwangyonhaedoktang plus hwangyon, Chongwisan, and Ssangbaksan showed remarkably potent antibacterial activity.

• Preventive Nutrition and Food Science
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• v.14 no.2
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• pp.162-167
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• 2009

Various fermented foods were screened in search of food-grade bacteria that produce bacteriocins active against Gram-negative pathogens. An isolate from a mold-ripened cheese presented antibacterial activity against Gram-positive and Gram-negative bacteria. The most active isolate was identified as Lactobacillus casei by a biochemical method, ribotyping, and membrane lipid analysis, and was designated as OSY-LB6A. The cell extracts of the isolate showed inhibition against Escherichia coli p220, E. coli O157, Salmonella enerica serovar Enteritidis, Salmonella Typhimurium, and Listeria monocytogenes. The antibacterial nature of the cell extract from the isolate was confirmed by eliminating the inhibitory effects of acid, hydrogen peroxide, and lytic bacteriophages. The culture supernatant and cell extract retained antibacterial activity after heating at $60{\sim}100^{\circ}C$ for $10{\sim}20$ min. The activity of the cell extract from Lb. casei was eliminated by pronase and lipase. Finally, the cell extract showed a bactericidal mode of action against E. coli in phosphate buffer solution, but it was bacteriostatic in broth medium and food extracts.

• Korean Journal of Veterinary Research
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• v.33 no.1
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• pp.53-60
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• 1993

Anaerobic bacteria are suggested to be potential source for new antibiotics. In order to search for antibiotics from domestic origin, we collected 800 soil samples across Korean locations and could isolate as many as 989 anaerobic strains. Among them 10, strains were found to have good producing capacity of antibiotics. An anaerobe was finally selected due to secreting antibiotics having high antimicrobial activity towards multiple resistant microorganism(E coli JM 83) transformed by genetic engineering technique. Its morphological, physiological and biochemical charateristics were investigated, together with antimicrobial spectrum therefrom. On antimicrobial spectrum study, substance secreted from this strain, had no activities to fungus and yeast. The selected strain showed G(+) and coccal shape, on Gram, staining and electron scanning microscopy, respectively. Biochemically this strain utilized glucose, fructose lactose, sucrose, but did not arabinose, cellulose, rhamnose, sorbitol, trehalose, mannitol. Catalase test showed negative property. Optimal growth temperature was $37^{\circ}C$. The results obtained above suggest this strain Streptococcus faecium subspp. and we named it Streptococcus sp. An-21-1.