• International Journal of Oral Biology
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• v.41 no.4
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• pp.199-208
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• 2016

The aim of this study was to identify the non-Aggregatibacter actinomycetemcomitans bacteria grown on the tryptic soy-serum-bacitracin-vancomycin (TSBV) medium, an A. actinomycetemcomitans selective medium. A total of 82 unidentified bacterial isolates from the oral cavities of a Korean population were kindly provide by the Korean Collection for Oral Microbiology. All the clinical isolates were grown on TSBV medium and bacterial DNA purified from each isolate was subjected to PCR with universal primers specific for bacterial 16S rRNA genes (16S rDNAs) sequence. The each bacterial 16S rDNA was amplified by PCR and the nucleotide sequences of it was determined by the dideoxynucleotide chain termination method. They were identified by 16S rDNA sequence comparison method at the specie-level. The data showed that Neisseria spp. (42 strains), Fusobacterium spp. (10 strains), Capnocytophaga spp. (8 strains), Propionibacterium acnes (5 strains), Aggregatibacter aprophilus (4 strains), Campylobacter spp. (5 strains), Veillonella dispar (3 strains), Streptococcus sp. (1 strain), Haemophilus parainfluenzae (1 strain), Leptotrichia wadei (1 strain), Morococcus sp./Neisseria sp. (1 strain), and Staphylococcus sp. (1 strain) were identified. These results could be used to develop a new A. actinomycetemcomitans-selective medium which is more effective than the TSBV medium in future studies.

• Journal of Periodontal and Implant Science
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• v.25 no.3
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• pp.487-496
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• 1995

Previous studies have demonstrated that not all A. actinomycetemcomitans produced significant level of leukotoxic factor and its leukotoxicity have associated with serotype and genetic variation. Our aim was to investigate on the interrelationship between serotype and leukotoxicity of an A. actinomycetemcomitans consisting of 13 clinically well characterized. Korean isolates and to evaluate if particular virulent clonal types of A. actinomycetemcomitans are associated with periodontal disease. For this study, 13 strains of A. actinomycetemcomitans from 6 patients with periodontal disease were isolated and identified by using a selective medium(tryptic soy agar supplemented with 10% serum, $75{\mu}g$ of bacitracin and $5{\mu}g$ of vancomycin per ml) in 10% C02 incubator for 3days with routine Gram staining, colony morphology and biochemical test..For serotyping, antisera were prepared from reference strains of 5 serotypes. (ATCC 29523,Y4, SUNY aB 67, IDH 781, IDH 1705) and then ammonium sulfate precipitation, immunoabsorption and indirect immunofluoroscent procedures were done. For analysis of leukotoxicity, sonic extract of A. actinomycetemcomitans exposed to PMN, and trypan blue was stained for counting the cell viability. Finally Southern blot analyses of genomic DNA digested with the restriction enzyme Tag I was done and the Southern blots were hybridized with the 530bp fragment, termed delta 530, originating from the ltx promoter of strain 652 and deleted from strain JP2. Also ltxA-3.1 and SC2 probe from strain JP2 were hybridized with genomic DNA fragments. Results reveal that strains isolated showed approximately equal proportions of 3 serotypes(b, d, e) and serotype b was not detected. 2 patients harbored 2 different serotypes in the same disease site. The prevalence of leukotoxic strain was 23% and there was no relationship between serotype, leukotoxicity and clinical observations. Especially virulent clonal types of Actinobacillus actinomycetemcomitan (JP2 strain) could not found. Further studies are necessary on the genetic polymorphism of leukotoxin and its relations to clinical status.

• The Journal of the Korean Society for Microbiology
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• v.21 no.4
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• pp.487-501
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• 1986

Previous studies from our laboratory suggested that Korean LJP patients might habor A. actinomycetemcomitans of different serotype from Caucasian LJP patients in whom serotype b was predominant. In order to observe the prevalence and serotype distribution of A. actinomycetemcomitans in localized juvenile periodontitis patients and to evaluate leukotoxic activity of oral isolates, this study was performed. A. actinomycetemcomitans was isolated by using a selective medium(tryptic soy agar supplemented with 10% serum, $75{\mu}g$ of bacitracin and $5{\mu}g$ of vancomycin per ml). Using immunoabsorbed, ammonium sulfate-fractionated serotype-specific antisera, a total of 69 strains were serologically categorized by ELISA. Leukotoxicity was monitored biochemically by measuring lactate dehydrogenase indicator of cell viability in culture supernatant of PMNL plus viable A. actinomycetemcomitans mixture. The results were as follows: 1. A. actinomycetemcomitans was detected in 75% of 16 LJP patients, and 71% in the LJP lesions and 6% in the control sites. 2. Presence or absence of A. actinomycetemcomitans in the sampled disease sites has no in fluence on clinical measurements. 3. Three serotypes were approximately equally distributed in overall 9 patients. Three patients harbored 2 different serotypes of A. actinomycetemcomitans in the same disease site or different disease sites. 4. The proportion of leukotoxic oral isolates was 22% of a total of 46 strains and the prevalence was 69% in 13 sampled sites. The same disease site could harbor both leukotoxic and nonleukotoxic strains. 5. Distribution of leukotoxic strains in 3 serotypes were not different.