Development of strain-specific polymerase chain reaction primers to detect Fusobacterium hwasookii strains

Yun Kyong Lim,Joong-Ki Kook 대한구강생물학회 2021 International Journal of Oral Biology Vol.46 No.4

This study aimed to develop strain-specific polymerase chain reaction (PCR) primers to detect Fusobacterium hwasookii KCOM 1249T, F. hwasookii KCOM 1253, F. hwasookii KCOM 1256, F. hwasookii KCOM 1258, and F. hwasookii KCOM 1268 on the basis of nucleotide sequences of a gene specific to each strain. The unique genes for each F. hwasookii strain were determined on the basis of their genome sequences using Roary. The strain-specific PCR primers based on each strain-specific gene were designed using PrimerSelect. The specificity of each PCR primer was determined using the genomic DNA of the 5 F. hwasookii strains and 25 strains of oral bacterial species. The detection limit and sensitivity of each strain-specific PCR primer pair were determined using the genomic DNA of each target strain. The results showed that the strain-specific PCR primers correspond to F. hwasookii KCOM 1249T, F. hwasookii KCOM 1253, F. hwasookii KCOM 1258, F. hwasookii KCOM 1256/F. nucleatum subsp. polymorphum KCOM 1260, or F. hwasookii KCOM 1268/Fusobacterium sp. oral taxon 203 were developed. The detection limits of these strain-specific PCR primers ranged from 0.2 to 2 ng of genomic DNA for each target strain. The results suggest that these strain-specific PCR primers are valuable in quality control for detecting specific F. hwasookii strains. gingivalis is a well-established periodontal pathogen, and tissue factor (TF) is a key initiator of the coagulation cascade. In this context, P. gingivalis has been reported to enhance TF expression in human endothelial cells. The present study investigated the underlying mechanisms of TF induction by P. gingivalis in human umbilical vein endothelial cells. P. gingivalis increased TF expression in a dose- and time-dependent manner. Not only live bacteria but also glutaraldehyde-fixed bacteria increased TF expression to the same extent. However, sonicates of P. gingivalis did not induce TF expression. Cytochalasin D and SMIFH2, which are inhibitors of actin polymerization and actin nucleation, respectively, inhibited the TF expression induced by P. gingivalis . Finally, TF production was decreased or increased in the presence of various signaling inhibitors, including mitogen-activated protein kinases. These results suggest that P. gingivalis induces endothelial TF expression by a bacterial internalization-dependent mechanism and through diverse signal transduction mechanisms. solutions. Tertiary dentin is formed when odontoblasts are directly affected by various stimuli. Recent preclinical studies have reported that stimulation of the Wnt/β-catenin signaling pathway could facilitate the formation of reparative dentin and thereby aid in the structural and functional development of the tertiary dentin. A range of signaling pathways, including the Wnt/β-catenin pathway, is activated when dental tissues are damaged and the pulp is exposed. The application of small molecules for dentin regeneration has been suggested as a drug repositioning approach. This study reviews the role of Wnt signaling in tooth formation, particularly dentin formation and dentin regeneration. In addition, the application of the drug repositioning strategy to facilitate the development of new drugs for dentin regeneration has been discussed in this study.

Strain-specific Detection of Kimchi Starter Leuconostoc mesenteroides WiKim33 using Multiplex PCR

Lee, Moeun,Song, Jung Hee,Park, Ji Min,Chang, Ji Yoon Korean Society of Food Culture 2019 韓國食生活文化學會誌 Vol.34 No.2

Leuconostoc spp. are generally utilized as kimchi starters, because these strains are expected to have beneficial effects on kimchi fermentation, including improvement of sensory characteristics. Here, we developed a detection method for verifying the presence of the kimchi starter Leuconostoc mesenteroides WiKim33, which is used for control of kimchi fermentation. A primer set for multiplex polymerase chain reaction was designed based on the nucleotide sequence of the plasmids in strain WiKim33, and their specificity was validated against 45 different strains of Leuconostoc spp. and 30 other strains. Furthermore, the starter strain consistently tested positive, regardless of the presence of other bacterial species in starter kimchi during the fermentation period. Our findings showed that application of a strain-specific primer set for strain WiKim33 presented a rapid, sensitive, and specific method for detection of this kimchi starter strain during natural kimchi fermentation.

Strain-specific Detection of Kimchi Starter Leuconostoc mesenteroides WiKim33 using Multiplex PCR

Moeun Lee,Jung Hee Song,Ji Min Park,Ji Yoon Chang 한국식생활문화학회 2019 韓國食生活文化學會誌 Vol.34 No.2

Leuconostoc spp. are generally utilized as kimchi starters, because these strains are expected to have beneficial effects on kimchi fermentation, including improvement of sensory characteristics. Here, we developed a detection method for verifying the presence of the kimchi starter Leuconostoc mesenteroides WiKim33, which is used for control of kimchi fermentation. A primer set for multiplex polymerase chain reaction was designed based on the nucleotide sequence of the plasmids in strain WiKim33, and their specificity was validated against 45 different strains of Leuconostoc spp. and 30 other strains. Furthermore, the starter strain consistently tested positive, regardless of the presence of other bacterial species in starter kimchi during the fermentation period. Our findings showed that application of a strain-specific primer set for strain WiKim33 presented a rapid, sensitive, and specific method for detection of this kimchi starter strain during natural kimchi fermentation.

Strain-specific Detection of Kimchi Starter Leuconostoc mesenteroides WiKim33 using Multiplex PCR

이모은,송정희,박지민,장지윤 한국식생활문화학회 2019 韓國食生活文化學會誌 Vol.34 No.2

Leuconostoc spp. are generally utilized as kimchi starters, because these strains are expected to have beneficial effects on kimchi fermentation, including improvement of sensory characteristics. Here, we developed a detection method for verifying the presence of the kimchi starter Leuconostoc mesenteroides WiKim33, which is used for control of kimchi fermentation. A primer set for multiplex polymerase chain reaction was designed based on the nucleotide sequence of the plasmids in strain WiKim33, and their specificity was validated against 45 different strains of Leuconostoc spp. and 30 other strains. Furthermore, the starter strain consistently tested positive, regardless of the presence of other bacterial species in starter kimchi during the fermentation period. Our findings showed that application of a strain-specific primer set for strain WiKim33 presented a rapid, sensitive, and specific method for detection of this kimchi starter strain during natural kimchi fermentation.

Development of Strain-specific PCR Primers Based on a DNA Probe Fu12 for the Identification of Fusobacterium nucleatum subsp. nucleatum ATCC $25586^T$

Kim Hwa-Sook,Song Soo Keun,Yoo So Young,Jin Dong Chun,Shin Hwan Seon,Lim Chae Kwang,Kim Myong Soo,Kim Jin-Soo,Choe Son-Jin,Kook Joong-Ki The Microbiological Society of Korea 2005 The journal of microbiology Vol.43 No.4

The objective of this study was to assess the strain-specificity of a DNA probe, Fu12, for Fusobacterium nucleatum subsp. nucleatum ATCC $25586^T$ (F. nucleatum ATCC $25586^T$), and to develop sets of strain-specific polymerase chain reaction (PCR) primers. Strain-specificity was tested against 16 strains of F. nucleatum and 3 strains of distinct Fusobacterium species. Southern blot hybridization revealed that the Fu12 reacted exclusively with the HindIII-digested genomic DNA of F. nucleatum ATCC $25586^T$. The results of PCR revealed that three pairs of PCR primers, based on the nucleotide sequence of Fu12, generated the strain-specific amplicons from F. nucleatum ATCC $25586^T$. These results suggest that the DNA probe Fu12 and the three pairs of PCR primers could be useful in the identification of F. nucleatum ATCC $25586^T$, especially with regard to the determination of the authenticity of the strain.

Development of Strain-specific PCR Primers Based on a DNA Probe Fu12 for the Identification of Fusobacterium nucleatum subsp. nucleatum ATCC 25586T

Hwa-Sook Kim,Soo Keun Song,유소영,Dong Chun Jin,Hwan Seon Shin,Chae Kwang Lim,김명수,김진수,Son-Jin Choe,국중기 한국미생물학회 2005 The journal of microbiology Vol.43 No.4

The objective of this study was to assess the strain-specificity of a DNA probe, Fu12, for Fusobacterium nucleatum subsp. nucleatum ATCC 25586T (F. nucleatum ATCC 25586T), and to develop sets of strainspecific polymerase chain reaction (PCR) primers. Strain-specificity was tested against 16 strains of F. nucleatum and 3 strains of distinct Fusobacterium species. Southern blot hybridization revealed that the Fu12 reacted exclusively with the HindIII-digested genomic DNA of F. nucleatum ATCC 25586T. The results of PCR revealed that three pairs of PCR primers, based on the nucleotide sequence of Fu12, generated the strain-specific amplicons from F. nucleatum ATCC 25586T. These results suggest that the DNA probe Fu12 and the three pairs of PCR primers could be useful in the identification of F. nucleatum ATCC 25586T, especially with regard to the determination of the authenticity of the strain.

국내 연안산 질소고정 단세포 남세균 종주의 광생물학적 수소생산력

박종우,명금옥,이원호 한국수소및신에너지학회 2010 한국수소 및 신에너지학회논문집 Vol.21 No.2

Photobiological hydrogen production by nitrogen-fixing unicellular cyanobacteria has long been considered to be an environmentally sound and very promising method for the future supply of renewable clean energy. We tried to find out the optimum cell concentration for H2 production in each of the two new Korean nitrogen-fixing unicellular cyanobacterial strains to compare with Synechococcus sp. strain Miami BG043511. The two Korean strains, Cyanothece sp. KNU CB MAL-031 and KNU CB MAL-058, were isolated from Korean west coasts. Cell concentrations up to 17 billion cells ml-1 were applied to the tests. High cell concentration over 15 billion cells ml-1 resulted in drastically reduced H2 production in all the three strains. The two domestic strains, however, produced 2-3 time more hydrogen than Synechococcus sp. Miami BG043511 at cell concentrations of 5-10 billion cells ml-1. At lower cell concentrations than 2 billion cells ml-1, MAL-031 exhibited highest H2 production followed by Miami BG043511, with far less production in MAL-058. Present result suggests that Cyanothece sp. MAL-CB031 might be one of the ideal nitrogen-fixing unicellular cyanobacterial strains for the photobiological hydrogen production.

한국 연안산 질소고정 단세포 남세균 종주의 최적 성장 및 수소생산 온도

박종우,김형섭,이원호 한국수소및신에너지학회 2013 한국수소 및 신에너지학회논문집 Vol.24 No.1

Photobiological hydrogen production by nitrogen-fixing unicellular cyanobacteria has long been considered to be an environmentally sound and very promising method for the future supply of renewable clean energy. Using six Korean nitrogen-fixing unicellular cyanobacterial strains and the Synechococcus sp. strain Miami BG043511 we performed cultivation experiments to find out the strain-specific optimal temperature for population growth and H2 production. Under 20℃ the population growth of all the tested strains was significantly retarded in contrasts to the faster and higher growth under 25, 30 or 35℃. The highest growth rates in all the 7 strains were measured under 30℃ while the maximal biomass yields were under 30℃ (strains CB-MAL 026, 054, and 055) or 35℃ (strains 002, 031, 058, and Miami BG043511). The difference between the maximal biomass yields at 30℃ and 35℃ was not greater than 10%. The quantity of photobiologically produced H2 was only slight larger under 35℃ than that under 20℃. Our result may suggest a two-step process of H2 production which includes rapid and sizable production of biomass at 30℃ and the following high H2 production at 20℃ by the test strains of marine nitrogen-fixing unicellular cyanobacteria.

Strain-specific PCR Primers for the Detection of Prevotella intermedia ATCC 49046

국중기 KOREAN ACADAMY OF ORAL BIOLOGY 2011 International Journal of Oral Biology Vol.36 No.2

The aim of this study was to develop Prevotella intermedia ATCC 49046-specific PCR primers designed based on the nucleotide sequence of a DNA probe Pig28. The strainspecificity of the PCR primers, Pig28-F1/Pig28-R1, was confirmed with 9 strains of P. intermedia and 25 strains (15 species) of Prevotella species. The detection limit of the PCR primers was 2 pg of the purified genomic DNA of P. intermedia ATCC 49046. These PCR primers were found to be useful for identifying P. intermedia ATCC 49046, particularly for determining the authenticity of the strain.

단세포성 해양남세균 종주를 이용한 광생물학적 수소생산 기술

박종우,김재만,이원호,Park, Jong-Woo,Kim, Jae-Man,Yih, Won-Ho 한국해양학회 2009 바다 Vol.14 No.1

광생물학적 수소생산 잠재력을 가진 다양한 미소생물 가운데, 남세균은 21세기의 수소경제 시대에 적합한 생물군으로 오랫동안 알려져 왔다. 광생물학적으로 수소에너지를 생산하게 될 경우, 해양 단세포성 질소고정 남세균은 남세균류의 하부 분류군들 가운데 가장이상적인 종류의 하나로 평가되고 있다. 단세포성 질소고정 남세균을 이용한 수소생산 기술을 개발하기 위해 반드시 고려해야 할 3가지 사항은 1) 자연계에 존재하는 최우수 수소생산 종주의 확립 2) 광생물학적 수소생산을 뒷받침하는 종주-특이적 최적조건의 탐색 3) 유전학적 방법을 이용한 수소생산 종주의 개량 등이다. 본고에서는 광생물학적 수소생산기술의 상업화를 향한 최근의 연구 개발 추세를 돌아보고, 해양 단세포성 남세균 종주를 이용한 광생물학적 수소생산 기술 분야에서 한국의 세계선도적 지위 확보를 위해서는 향후 10-15년간 집중적인 연구 개발이 절실함을 제안하고자 한다. Among various microscopic organisms producing photobiological hydrogen, cyanobacteria have long been recognized as the promising biological agents for hydrogen economy in 21 century. For photobiological production of hydrogen energy, marine unicellular $N_2$-fixing cyanobacteria have been evaluated as an ideal subgroup of Cyanophyceae. To develope the hydrogen production technology using unicellular $N_2$-fixing cyanobacteria, 3 important factors are pre-requisite: 1) isolation of the best strain from marine natural environment, 2) exploration on the strain-specific optimal conditions for the photobiological hydrogen production, and finally 3) application of the molecular genetic tools to improve the natural ability of the strain to produce hydrogen. Here we reviewed the recent research & development to commercialize photobiological hydrogen production technology, and suggest that intensive R&D during next 10-15 years should be imperative for the future Korean initiatives in the field of the photobiological hydrogen production technology using photosynthetic marine unicellular cyanobacterial strains.