ABSTRACT
The aim of this study was to isolate and detect microorganisms of root-filled teeth associated with periradicular lesions. Specimens were sampled from patients undergoing root canal retreatment. The bacteria were characterized by morphologic and biochemical analysis and by 16S rRNA gene sequencing. Microorganisms were detected in 10 of 18 teeth. The majority of positive samples revealed a mixed culture of 2-8 species. In 2 teeth Enterococcus faecalis was the only detected species. For the first time Vagococcus fluvialis was detected in root canals. Solobacterium moorei and Fusobacterium nucleatum were the most prevalent species. Presence of F. nucleatum was associated with the presence of S. moorei in 5 of 7 cases. In all teeth with Parvimonas micra and Dialister invisus, F. nucleatum and S. moorei were found. Moreover, members of additional different genera were detected delivering bacterial compositions that have been not described yet.
Subject(s)
Dental Pulp Cavity/microbiology , Periapical Periodontitis/microbiology , Tooth, Nonvital/microbiology , Adult , Aged , Bacterial Typing Techniques , Colony Count, Microbial , DNA, Bacterial/analysis , Fusobacterium nucleatum/isolation & purification , Gram-Positive Asporogenous Rods/isolation & purification , Gram-Positive Asporogenous Rods/pathogenicity , Gram-Positive Cocci/isolation & purification , Gram-Positive Cocci/pathogenicity , Humans , Middle Aged , Periapical Periodontitis/therapy , Polymerase Chain Reaction , RNA, Ribosomal, 16S/genetics , Retreatment , Root Canal TherapyABSTRACT
Renibacterium salmoninarum is a facultative intracellular pathogen often found in host phagocytes where it appears to successfully avoid the host fish's immunological defenses. The objective of this investigation was to determine if soluble extracellular protein (ECP) produced by R. salmoninarum may contribute to the immunomodulation in bacterial kidney disease (BKD) via inhibition of phagocyte respiratory burst and/or phagocytosis mechanisms. Splenic cells from adult brook trout (Salvelinus fontinalis) were incubated with two different concentrations of ECP (0.1 mg/ml and 1.0 mg/ml) and viable R. salmoninarum. Splenic cell cultures were evaluated for respiratory burst activity via flow cytometry with the dichlorofluorescin diacetate (DCF-DA) assay and for phagocytosis via light microscopic assessment of microsphere engulfment. Respiratory burst activity was significantly inhibited in all treatment groups as compared to untreated fish, while no differences were noted in phagocytic activity.
Subject(s)
Bacterial Proteins/toxicity , Gram-Positive Asporogenous Rods/immunology , Gram-Positive Asporogenous Rods/pathogenicity , Phagocytes/drug effects , Phagocytes/immunology , Trout/immunology , Animals , Fish Diseases/etiology , Fish Diseases/immunology , Fish Diseases/microbiology , Fluoresceins , Gram-Positive Bacterial Infections/etiology , Gram-Positive Bacterial Infections/immunology , Gram-Positive Bacterial Infections/veterinary , Kidney Diseases/etiology , Kidney Diseases/immunology , Kidney Diseases/veterinary , Microspheres , Phagocytes/metabolism , Phagocytosis/drug effects , Respiratory Burst/drug effects , Spleen/cytology , Spleen/immunology , Spleen/metabolism , Trout/metabolismABSTRACT
Two homogeneous proteins active in vitro against the bacterial pathogen Clavibacter michiganensis subsp. sepedonicus were obtained from a crude cell-wall preparation from the leaves of Columbia wild-type Arabidopsis. The N-terminal amino acid sequences of these proteins allowed their identification as lipid transfer proteins (LTP-a1, LTP-a2); the LTP1-a1 sequence was identical to that deduced from a previously described cDNA (EMBL M80566) and LTP-a2 was quite divergent (44% identical positions). These proteins were not detected in the cytoplasmic fraction by Western-blot analysis. Proteins LTP-s1 and LTP-s2 were similarly obtained from spinach leaves; LTP-s1 was 91% identical to a previously purified spinach LTP (Swiss Prot P10976), and LTP-s2 was moderately divergent (71% identical positions). About 1/3 of the total LTPs were detected in the cytoplasmic fraction from spinach by Western-blot analysis. Concentrations of these proteins causing 50% inhibition (EC-50) were in the 0.1-1 microM range for the bacterial pathogens C. michiganensis and Pseudomonas solanacearum and close to 10 microM for the fungal pathogen Fusarium solani.
Subject(s)
Arabidopsis/metabolism , Carrier Proteins/isolation & purification , Carrier Proteins/toxicity , Gram-Positive Asporogenous Rods/drug effects , Vegetables/metabolism , Amino Acid Sequence , Antigens, Plant , Arabidopsis/microbiology , Bacteria, Aerobic/drug effects , Bacteria, Aerobic/pathogenicity , DNA, Complementary , Fusarium/drug effects , Fusarium/pathogenicity , Gram-Positive Asporogenous Rods/pathogenicity , Microbial Sensitivity Tests , Molecular Sequence Data , Molecular Weight , Plant Proteins , Pseudomonas/drug effects , Pseudomonas/pathogenicity , Sequence Homology, Amino Acid , Vegetables/microbiologySubject(s)
Gram-Positive Asporogenous Rods/pathogenicity , Gram-Positive Bacterial Infections/microbiology , Gram-Positive Cocci/pathogenicity , Lactates/biosynthesis , Gram-Positive Asporogenous Rods/isolation & purification , Gram-Positive Asporogenous Rods/metabolism , Gram-Positive Cocci/isolation & purification , Gram-Positive Cocci/metabolism , Humans , Lactic AcidABSTRACT
A gene encoding haemolytic activity from Renibacterium salmoninarum (strain PPD) was cloned into Escherichia coli using the cosmid vector pHC79, and subsequently subcloned on a 1.6 kbp SAlI fragment into pBR328. Southern blot hybridisation revealed that a homologous sequence is found in other strains of R. salmoninarum.
