Isolation and molecular identification of Fusobacterium nucleatum from Nigerian patients with oro-facial infections.

BACKGROUND: Fusobacterium nucleatum is one of the most common anaerobic bacteria present in the oral cavity and is often isolated from infections involving other body sites. OBJECTIVE: To characterise F. nucleatum strains from patients attending a teaching hospital in Nigeria in order to provide information on the methods for accurate identification of anaerobes in clinical specimen. METHODS: Fusobacterium nucleatum specie from 50 patients presenting with oro-facial infections were studied by culture on Fusobacterium selective agar and fastidious anaerobe agar. The isolates were characterised based on colonial morphology, microscopy, lipase production, susceptibility to kanamycin and colistin and resistance to vancomycin. Biochemical tests were performed using a commercial test kit. The identity of the isolates was confirmed based on molecular characterization performed using polymerase chain reaction (PCR) analysis. RESULTS: Forty-eight (96%) F. nucleatum isolates were obtained from the 50 patients by culture and all the isolates were identified by colonial appearance and microscopy based on their unique spindle shape with tapered ends. Only 26 (54.2%) of the 48 isolates were identified by commercial API 20A test kit while PCR confirmed the identity of all the isolates. CONCLUSION: Anaerobes are involved in human infections and their study is quite cumbersome due to tedious nature and high cost of the techniques involved. Cultural method is reliable in the isolation and identification of F. nucleatum species. PCR is a rapid and simple method that can complement the phenotypic identification of anaerobes and would assist in their full identification.

Effect of Psidium cattleianum leaf extract on Streptococcus mutans viability, protein expression and acid production.

Plants naturally produce secondary metabolites that can be used as antimicrobials. The aim of this study was to assess the effects of Psidium cattleianum leaf extract on Streptococcus mutans. The extract (100%) was obtained by decoction of 100 g of leaves in 600 ml of deionized water. To assess killing, S. mutans biofilms were treated with water (negative control) or various extract dilutions [100, 50, 25% (v/v) in water] for 5 or 60 min. To evaluate the effect on protein expression, biofilms were exposed to water or 1.6% (v/v) extract for 120 min, proteins were extracted and submitted to 2-dimensional difference gel electrophoresis. Differentially expressed proteins were identified by mass spectrometry. The effect of 1.6% (v/v) extract on acid production was determined by pH measurements and compared to a water control. Viability was similar after 5 min of treatment with the 100% extract or 60 min with the 50% extract (about 0.03% survival). There were no differences in viability between the biofilms exposed to the 25 or 50% extract after 60 min of treatment (about 0.02% survival). Treatment with the 1.6% extract significantly changed protein expression. The abundance of 24 spots was decreased compared to water (p < 0.05). The extract significantly inhibited acid production (p < 0.05). It is concluded that P. cattleianum leaf extract kills S. mutans grown in biofilms when applied at high concentrations. At low concentrations it inhibits S. mutans acid production and reduces the expression of proteins involved in general metabolism, glycolysis and lactic acid production.

Influence of antimicrobial subinhibitory concentrations on hemolytic activity and bacteriocin-like substances in oral Fusobacterium nucleatum.

Fusobacterium nucleatum is considered for its role in colonization of initial and late microorganisms in dental plaque and for its coaggregation with other bacterial species. It is known that action of different antimicrobial substances may interfere with either virulence factors or with host-bacteria interaction. The goal of this study was to examine the influence of subinhibitory concentrations of chlorhexidine, triclosan, penicillin G and metronidazole on hemolytic activity and bacteriocin-like substance production of oral F. nucleatum. A high resistance to penicillin G was observed and 63% of the isolates were beta-lactamase positive. All the tested isolates were susceptible to metronidazole. F. nucleatum isolates grown with or without antimicrobials were alpha-hemolytics. Bacteriocin-like substance production was increased in isolates grown with penicillin G. Impaired production of hemolytic or antagonic substances can suggest a role in the regulation of oral microbiota.

Oral species of Fusobacterium from human and environmental samples.

PURPOSE: The aim of this study was the characterization and identification of oral Fusobacterium in patients with and without periodontal disease, and from spittoons and air-water syringes. The antimicrobial susceptibility of this bacterium was evaluated. METHOD: Subgingival samples were taken using sterilized absorbent paper points. Spittoon samples were collected using sterile swabs around the drain area with shut off, and air-water syringe samples by washing the tip with Ringer solution. Samples were transferred in tubes under CO2 flux. Diluted samples were inoculated on to Omata and Disraely agar and blood agar plates, which were incubated in anaerobiosis, at 37 degrees C, for 4 days. Bacterial species were identified biochemically. MIC was determined using an agar dilution method. RESULTS: Periodontal patients, healthy subjects, spittoons and air-water syringes were 80%, 67.6%, 37.8% and 3.3% positive to Fusobacterium, respectively. Clindamycin, imipenem, lincomycin, metronidazole and tetracycline were active against all human and environmental isolates. Eighteen isolates resistant to ampicillin or penicillin G produced beta-lactamases. The presence of human oral bacteria in items of dental equipment supports the hypothesis that such equipment may serve as a vehicle for the transmission of pathogenic organisms. CONCLUSION: Pieces of dental equipment may serve as a vehicle for the transmission of oral pathogenic organisms.