A multiplex PCR assay was developed for the simultaneous detection of the etiologic agents associated with porcine proliferative enteropathies (PPE), swine dysentery (SD)and porcine salmonellosis (PS)in a single reaction using DNA from swine intestinal samples. Single and multiplex PCR amplification of DNA from Lawsonia intracellularis, Salmonella typhimurium and Brachyspira hyodysenteriae with each primer set produced fragments of the predicted size without any nonspecific amplification, 210-bp, 298-bp and 403-bp bands, respectively. The single PCR assay could detect as little as 100 pg of purified DNA of S. typhimurium and L. intracellularis, and 50 pg of B.hyodysenteriae, respectively. However, multiplex PCR turned out to be 10 times lower sensitivity with S. typhimurium compared with single PCR. With 23 swine intestinal specimens suspected of having PPE, SD and/or PS, the multiplex PCR assay showed identical results with conventional methods except one. In conclusion, this multiplex PCR is a feasible alternative to standard diagnostic methods for detection of L. intracellularis, B. hyodysenteriae and Salmonella spp. from swine intestinal specimens.
Animals ; Desulfovibrionaceae Infections/microbiology/veterinary ; Intestines/microbiology ; Lawsonia Bacteria/*isolation&purification ; Polymerase Chain Reaction/*methods/veterinary ; Salmonella/*isolation&purification ; Salmonella Infections, Animal/diagnosis ; Sensitivity and Specificity ; Spirochaetales/*isolation&purification ; Spirochaetales Infections/microbiology/veterinary ; Swine ; Swine Diseases/*diagnosis/*microbiology

Using three reference strains of Brachyspira hyodysenteriae (B204, B234, B169), one B. pilosicoli (P43/6/78), one B. murdochii (56-150), one B. intermedia (PWS/A), one B. innocens (B256) and ten Korean isolates, PCR-RFLP analysis of DNA encoding 23S rRNA was performed to establish a rapid and accurate method for characterizing porcine intestinal spirochetes. Consequently, B. hyodysenteriae and B. pilosicoli revealed different restriction patterns; however, the other three species shared the same pattern. These findings are not consistent with a prior report. Differences in 23S rRNA gene sequences, between two B. murdochii strains, 56-150 and 155-20, were observed. These results indicate that 23S rRNA PCR-RFLP could be used as an identification method for pathogenic Brachyspira spp. (B. hyodysenteriae and B. pilosicoli) as well as an epidemiological tool for characterizing spirochetes isolated from swine.
Animals ; DNA, Bacterial/genetics ; Dysentery, Bacillary/diagnosis/microbiology/*veterinary ; Korea ; Phylogeny ; Polymerase Chain Reaction/methods/*veterinary ; Polymorphism, Restriction Fragment Length ; RNA, Ribosomal, 23S/chemistry/*genetics ; Spirochaetales/*genetics/*isolation&purification ; Spirochaetales Infections/diagnosis/microbiology/*veterinary ; Swine ; Swine Diseases/diagnosis/*microbiology

This study was done to characterize diversity in 10 Brachyspira hyodysenteriae isolates in Korea. The isolates were compared with 14 well-characterized non-Korean strains of various Brachyspira species. All Korean isolates showed strong beta haemolysis and had blunt cell ends with 7~14 periplasmic flagella. They produced indole, and did not ferment fructose. They were alpha-glucosidase positive and alpha-galatosidase negative using the APIZYM kit. Using polyclonal antisera raised in rabbits against recognized serotypes, all isolates showed a strong reaction to B. hyodysenteriae antisera E, A and B. Using multilocus enzyme electrophoresis (MLEE) with 15 enzymes and 5 buffer systems, the Korean and non-Korean isolates were divided into 22 electrophoretic types (ETs) and 5 divisions (A, B, C, D and E). Division A corresponded to B. hyodysenteriae, B to B. innocens, C to B. intermedia, D to B. murdochii and E to B. pilosicoli. The 10 Korean isolates of B. hyodysenteriae were relatively diverse, being divided into 9 ETs within MLEE division A. They were all distinct from the non-Korean strains.
Animals ; Electrophoresis ; Genes, Bacterial ; Korea/epidemiology ; Rabbits ; Serotyping ; Serpulina hyodysenteriae/classification/genetics/*isolation&purification ; Spirochaetales Infections/*microbiology ; Swine/microbiology ; Swine Diseases/*microbiology ; Variation (Genetics)

OBJECTIVETo investigate the microbiological changes of subgingival microbials in patients with gingivitis and wearing fixed orthodontic appliances.

METHODS48 subjects (10 to 17 years old) with gingivitis, and wearing fixed orthodontic appliances, were divided randomly into three groups (placebo, NS and CH). Placebo group had normal saline mouthrinse; only and no oral hygiene instruction (OHI). The NS group had OHI and normal saline mouthrinse; The CH group had OHI and 0.12% chlorhexidine gluconate mouthrinse. Bacterial examinations were carried out on baseline, one week, one month and three months after scaling. The bacterial examination was carried out. The percentage of coccus, bacillus and spirochete was calculated.

RESULTSIn placebo group and NS group, the percentage of coccus increased up to the third examination then dropped down. The spirochete's percentage changed inversely. CH group maintained an increasing trend in coccus' percentage and decreasing trend in spirochete's percentage. The percentage changes of coccus and bacillus between placebo group and CH group are statistically significant (P < 0.05).

CONCLUSIONSDuring the three-month examination, the CH group had better microbiologic change than the other two groups.

Adolescent ; Child ; Chlorhexidine ; administration & dosage ; analogs & derivatives ; Female ; Gingivitis ; microbiology ; prevention & control ; Humans ; Male ; Malocclusion ; microbiology ; therapy ; Mouthwashes ; Orthodontic Appliances ; Orthodontics, Corrective ; adverse effects ; methods ; Periodontal Attachment Loss ; microbiology ; prevention & control ; Periodontal Diseases ; microbiology ; prevention & control ; Periodontal Pocket ; microbiology ; pathology ; Spirochaetales ; isolation & purification ; Spirochaetales Infections ; microbiology

The prevalence of Lawsonia intracellularis, Brachyspira hyodysenteriae and Salmonella spp. were investigated by multiplex PCR using fecal samples of pigs with diarrhea or a history of diarrhea. The overall herd prevalence of L. intracellularis, B. hyodysenteriae and Salmonella spp. were 46.5%, 37.2% and 51.1%, respectively. Also, the prevalence of L. intracellularis, B. hyodysenteriae and Salmonella spp. among all sampled pigs were 19.9%, 10.8% and 17.7%, respectively. Seventeen of 43 herds were positive with 2 enteric organisms, and 2 herds were positive with L. intracellularis, B. hyodysenteriae and Salmonella spp. simultaneously. It was notable that 11 of 12 herds with more than 2, 000 pigs were affected with Salmonella spp., and that only 2 of 12 the herds were affected with B. hyodysenteriae. This study suggested that herds positive for L. intracellularis, B. hyodysenteriae and Salmonella spp. were distributed throughout Korea, although the relationship among other pathogens such as viral or parasitic ones and/or with metabolic disorders was not determined.
Animals ; DNA, Bacterial/isolation&purification ; Desulfovibrionaceae Infections/epidemiology/microbiology/*veterinary ; Diarrhea/microbiology/veterinary ; Korea/epidemiology ; *Lawsonia Bacteria/isolation&purification ; Polymerase Chain Reaction/veterinary ; Prevalence ; *Salmonella/isolation&purification ; Salmonella Infections, Animal/*epidemiology/microbiology ; *Serpulina hyodysenteriae/isolation&purification ; Spirochaetales Infections/epidemiology/microbiology/*veterinary ; Swine ; Swine Diseases/*epidemiology/microbiology