• The Plant Pathology Journal
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• v.29 no.1
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• pp.59-66
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• 2013

Colonization studies previously performed with a green-fluorescent-protein, GFP, labeled derivative of Bacillus amyloliquefaciens FZB42 revealed that the bacterium behaved different in colonizing surfaces of plant roots of different species (Fan et al., 2012). In order to extend these studies and to elucidate which genes are crucial for root colonization, we applied targeted mutant strains to Arabidopsis seedlings. The fates of root colonization in mutant strains impaired in synthesis of alternative sigma factors, non-ribosomal synthesis of lipopeptides and polyketides, biofilm formation, swarming motility, and plant growth promoting activity were analyzed by confocal laser scanning microscopy. Whilst the wild-type strain heavily colonized surfaces of root tips and lateral roots, the mutant strains were impaired in their ability to colonize root tips and most of them were unable to colonize lateral roots. Ability to colonize plant roots is not only dependent on the ability to form biofilms or swarming motility. Six mutants, deficient in abrB-, sigH-, sigD-, nrfA-, yusV and RBAM017410, but not affected in biofilm formation, displayed significantly reduced root colonization. The nrfA- and yusV-mutant strains colonized border cells and, partly, root surfaces but did not colonize root tips or lateral roots.

• Journal of Pharmacopuncture
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• v.25 no.1
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• pp.37-45
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• 2022

Objectives: The quorum-sensing-inhibitory and anti-biofilm activities of the methanol extract of E. globulus leaves were determined against clinically isolated multidrug-resistant Pseudomonas aeruginosa. Methods: The preliminary anti-quorum-sensing (AQS) activity of eucalyptus was investigated against a biosensor strain Chromobacterium violaceum ATCC 12472 (CV12472) by using the agar well diffusion method. The effect of sub-minimum inhibitory concentrations (sub-MICs) of the methanol extract of eucalyptus on different quorum-sensing-regulated virulence factors, such as swarming motility, pyocyanin pigment, exopolysaccharide (EPS), and biofilm formation, against clinical isolates (CIs 2, 3, and 4) and reference PA01 of Pseudomonas aeruginosa were determined using the swarm diameter (mm)-measurement method, chloroform extraction method, phenol (5%)-sulphuric acid (concentrated) method, and the microtiter plate assay respectively, and the inhibition (%) in formation were calculated. Results: The preliminary AQS activity (violacein pigment inhibition) of eucalyptus was confirmed against Chromobacterium violaceum ATCC 12472 (CV12472). The eucalyptus extract also showed concentration-dependent inhibition (%) of swarming motility, pyocyanin pigment, EPS, and biofilm formation in different CIs and PA01 of P. aeruginosa. Conclusion: Our results revealed the effectiveness of the E. globulus extract for the regulation of quorum-sensing-dependent virulence factors and biofilm formation at a reduced dose (sub-MICs) and suggest that E. globulus may be a therapeutic agent for curing and controlling bacterial infection and thereby reducing the possibility of resistance development in pathogenic strains.

• Microbiology and Biotechnology Letters
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• v.43 no.4
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• pp.357-366
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• 2015

Quorum sensing (QS) is involved in the process of cell-to-cell communication and as a gene regulatory mechanism, which has been implicated in bacterial pathogenicity. Bacteria use this QS system to control a variety of physiological processes. In this study, pomegranate (Punica granatum L.) peel extract (PPE) was first screened for its ability to inhibit QS in bio-reporter strains (Chromobacterium violaceum and C. violaceum CV026). Next, the ability of PPE to inhibit swimming motility and biofilm formation was examined in Y. enterocolitica. Additionally, changes in the expression of specific genes involved in the synthesis of the N-acylhomoserine lactones (AHLs; yenI and yenR) and in the flagellar regulon (fliA, fleB and flhDC) were evaluated by reverse transcription (RT)-PCR. The results show that PPE specifically inhibited and reduced QS-controlled violacein production by 78.5% in C. violaceum CV026, and decreased QS-associated biofilm formation and swimming motility in Y. enterocolitica without significantly affecting bacterial growth. These inhibitory effects were also associated with the down-regulation of gene expression involved in the synthesis of AHLs and in motility. Our results suggest that PPE could be a potential therapeutic agent to prevent enteropathogens in humans, as well as highlight the need to further investigate the in vivo properties of PPE for clinical applications.

• Journal of Microbiology and Biotechnology
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• v.28 no.3
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• pp.454-464
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• 2018

Salivary microbiota alterations can correlate with dental caries development in children, and mechanisms mediating this association need to be studied in further detail. Our study explored salivary microbiota shifts in children and their association with the incidence of dental caries with dentine involvement. Salivary samples were collected from children with caries and their subsequently matched caries-free controls before and after caries development. The microbiota was analyzed by 16S rRNA gene-based high-throughput sequencing. The salivary microbiota was more diverse in caries-free subjects than in those with dental caries with dentine involvement (DC). Although both groups exhibited similar shifts in microbiota composition, an association with caries was found by function prediction. Analysis of potential microbiome functions revealed that Granulicatella, Streptococcus, Bulleidia, and Staphylococcus in the DC group could be associated with the bacterial invasion of epithelial cells, phosphotransferase system, and ${\text\tiny{D}}-alanine$ metabolism, whereas Neisseria, Lautropia, and Leptotrichia in caries-free subjects could be associated with bacterial motility protein genes, linoleic acid metabolism, and flavonoid biosynthesis, suggesting that functional differences in the salivary microbiota may be associated with caries formation. These results expand the current understanding of the functional significance of the salivary microbiome in caries development, and may facilitate the identification of novel biomarkers and treatment targets.

• Microbiology and Biotechnology Letters
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• v.34 no.4
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• pp.289-298
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• 2006

3',5'-cyclic adenosine monophosphate (cAMP) is an important molecule, which mediates diverse cellular processes. For example, it is involved in regulation of sugar uptake/catabolism, DNA replication, cell division, and motility in various acterial species. In addition, cAMP is one of the critical regulators for syntheses of virulence factors in many pathogenic bacteria. It is believed that cAMP acts as a signal for environmental changes as well as a regulatory factor for gene expressions. Therefore, intracellular concentration of cAMP is finely modulated by according to its rates of synthesis (by adenylate cyclase), excretion, and degradation (by cAMP phosphodiesterase). In the present review, we discuss the bacterial physiological characteristics governed by CAMP and the molecular mechanisms for gene regulation by cAMP. Furthermore, the effect of cAMP on phosphotransferase system is addressed.

• The Plant Pathology Journal
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• v.39 no.3
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• pp.235-244
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• 2023

Acidovorax citrulli (Ac) is a phytopathogenic bacterium that causes bacterial fruit blotch (BFB) in cucurbit crops, including watermelon. However, there are no effective methods to control this disease. YggS family pyridoxal phosphate-dependent enzyme acts as a coenzyme in all transamination reactions, but its function in Ac is poorly understood. Therefore, this study uses proteomic and phenotypic analyses to characterize the functions. The Ac strain lacking the YggS family pyridoxal phosphate-dependent enzyme, AcΔyppAc(EV), virulence was wholly eradicated in geminated seed inoculation and leaf infiltration. AcΔyppAc(EV) propagation was inhibited when exposed to L-homoserine but not pyridoxine. Wild-type and mutant growth were comparable in the liquid media but not in the solid media in the minimal condition. The comparative proteomic analysis revealed that YppAc is primarily involved in cell motility and wall/membrane/envelop biogenesis. In addition, AcΔyppAc(EV) reduced biofilm formation and twitching halo production, indicating that YppAc is involved in various cellular mechanisms and possesses pleiotropic effects. Therefore, this identified protein is a potential target for developing an efficient anti-virulence reagent to control BFB.

• The Plant Pathology Journal
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• v.37 no.6
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• pp.673-680
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• 2021

Acidovorax citrulli (Ac) is the causative agent of bacterial fruit blotch disease in watermelon. Since resistant cultivars have not yet been developed, the virulence factors/mechanisms of Ac need to be characterized. This study reports the functions of a putative pyridoxal phosphate-dependent aminotransferase (PpdaAc) that transfers amino groups to its substrates and uses pyridoxal phosphate as a coenzyme. It was observed that a ppdaAc knockout mutant had a significantly reduced virulence in watermelon when introduced via germinated-seed inoculation as well as leaf infiltration. Comparative proteomic analysis predicted the cellular mechanisms related to PpdaAc. Apart from causing virulence, the PpdaAc may have significant roles in energy production, cell membrane, motility, chemotaxis, post-translational modifications, and iron-related mechanisms. Therefore, it is postulated that PpdaAc may possess pleiotropic effects. These results provide new insights into the functions of a previously unidentified PpdaAc in Ac.

• Reproductive and Developmental Biology
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• v.36 no.3
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• pp.163-166
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• 2012

This study was conducted to determine the relationship between elapsed time after semen preservation on the changes of bacteria and semen quality. Semen was diluted with BTS(Beltsville Thawing Solution) extender without antibiotic for 7 days and sperm parameter and fertility were measured. Sperm motility was measured by CASA and total bacteria number was counted after 22~24 hr incubation from counting agar plate in which sperm dilute to $10{\sim}10^6$ in 0.9% saline solution and inoculate to agar. Acrosomal integrity was measured by Chlortetracycline (CTC) staining. CTC patterns were uniform fluorescence over the whole head (pattern F), characteristic of incapacitated acrosome-intact spermatozoa; fluorescence-free band in the post-acrosomal region (pattern B), characteristic of capacitated acrosome-intact spermatozoa; and almost no fluorescence over the whole head except for a thin band in the equatorial segment (pattern AR), characteristic of acrosome reacted spermatozoa. Total number of bacteria was significantly increased (p<0.0001) 3 days after preservation. Sperm motility, viability, and morphological abnormality on elapsed time after preservation were lower from 5 ($77.24{\pm}6.47$, p<0.001) and 7 days ($77.24{\pm}6.47$, p<0.001) after preservation compared to 1 ($15.71{\pm}7.18$) and 3 days($18.39{\pm}7.22$) after preservation, respectively. Sperm viability was significantly lower ($53.25{\pm}35.03$, p<0.0001) at 7 days after preservation. Morphological abnormality of sperm was lower (p<0.001) at 1 ($15.71{\pm}7.18$) and 3 ($18.39{\pm}7.22$) days compared to 5 ($21.84{\pm}7.91$) and 7 ($22.59{\pm}9.93$) days after preservation. Acrosomal integrity and capacitation rate (pattern F) were significantly lower (p<0.001) from 5 days after preservation. Based on the data we obtained from this study suggested that semen preserved more than 5 days without antibiotic would not recommend use for artificial insemination.

• Journal of Microbiology and Biotechnology
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• v.17 no.3
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• pp.420-427
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• 2007

We investigated the temperature effects on the virulence, development, reproduction, and otility of two Korean isolates of entomopathogenic nematodes, Steinernema glaseri Dongrae strain and S. longicaudum Nonsan strain. In addition, we studied the growth and virulence of their respective symbiotic bacterium, Xenorhabdus poinarii for S. glaseri and Xenorhabdus sp. for S. longicaudum, in an insect host at different temperatures. Insects infected with the nematode-bacterium complex or the symbiotic bacterium was placed at $13^{\circ}C,\;18^{\circ}C,\;24^{\circ}C,\;30^{\circ}C,\;or\;35^{\circ}C$ in the dark and the various parameters were monitored. Both nematode species caused mortality at all temperatures tested, with higher mortalities occurring at temperatures between $24^{\circ}C\;and\;30^{\circ}C$. However, S. longicaudum was better adapted to cold temperatures and caused higher mortality at $18^{\circ}C$ than S. glaseri. Both nematode species developed to adult at all temperatures, but no progeny production occurred at $13^{\circ}C\;or\;35^{\circ}C$. For S. glaseri, nematode progeny production was best at inocula levels above 20 infective juveniles/host at $24^{\circ}C\;and\;30^{\circ}C$, but for S. longicaudum, progeny production was generally better at $24^{\circ}C$. Steinernema glaseri showed the greatest motility at $30^{\circ}C$, whereas S. longicaudum showed good motility at $24^{\circ}C\;and\;30^{\circ}C$. Both bacterial species grew at all tested temperatures, but Xenorhabdus sp. was more virulent at low temperatures $(13^{\circ}C\;and\;18^{\circ}C)$ than X. poinarii.

• Molecules and Cells
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• v.28 no.4
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• pp.389-395
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• 2009

We previously observed that the transcription of some flagellar genes decreased in Salmonella Typhimurium tdcA mutant, which is a gene encoding the transcriptional activator of the tdc operon. Since flagella-mediated bacterial motility accelerates the invasion of Salmonella, we have examined the effect of tdcA mutation on the invasive ability as well as the flagellar biosynthesis in S. Typhimurium. A tdcA mutation caused defects in motility and formation of flagellin protein, FliC in S. Typhimurium. Invasion assays in the presence of a centrifugal force confirmed that the defect of flagellum synthesis decreases the ability of Salmonella to invade into cultured epithelial cells. In addition, we also found that the expression of Salmonella pathogenicity island 1 (SPI1) genes required for Salmonella invasion was down-regulated in the tdcA mutant because of the decreased expression of fliZ, a positive regulator of SPI1 transcriptional activator, hilA. Finally, the virulence of a S. Typhimurium tdcA mutant was attenuated compared to a wild type when administered orally. This study implies the role of tdcA in the invasion process of S. Typhimurium.