Hydroxyapatite is a calcium phosphate bioceramic that has been shown by many authors to be biocompatible with bioactive properties. It is widely accepted as the best synthetic material available for surgical use as a bone graft substitute. HA granules produced by AMREC-SIRIM from local materials underwent 5 types of sterilisation techniques with different ageing periods. Samples were tested for chemical and phase composition and microbial contamination before and after being sterilised. From the microbiological tests done, none of the unsterilised positive control yielded a positive culture. Results from X-Ray diffraction studies found that all the sterilisation techniques did not chemically degrade or structurally change the HA granules significantly.
Bacteriological Techniques ; Bone Substitutes/*analysis ; Calcium Phosphates/*analysis ; Materials Testing/*methods ; Sterilization/*methods ; X-Ray Diffraction

To seek a practical and inexpensive method to preserve gonococcal cultures, a few methods were compared. The following methods kept the cultures alive for only a short period of time: skim milk at -20 degrees C; tryptic soy broth with 15% glycerol at -20 degrees C; cystine tryptic agar at 35 degrees C. Most of the test cultures survived for more than 4 weeks in the following media: one half strength dextrose starch agar; one half strength dextrose starch agar with ferric nitrate; one half strength dextrose starch agar with antimicrobic CNV. Dextrose starch agar could be substituted by GC medium base with a slight modification. It is concluded that preservation of gonococci in one half strength dextrose starch agar with CNV, which produces less frequent contamination, is a practical method to maintain cultures for teaching and quality control.
Agar ; Bacteriological Techniques ; Culture Media* ; Glucose ; Neisseria gonorrhoeae* ; Preservation, Biological/methods* ; Starch

Clostridium cellulolyticum, as one of obligate anaerobic bacteria capable of secreting cellulosome, has not been efficiently cultured due to its strict requirement of growing conditions. In this study, culture conditions of C. cellulolyticum were optimized using response surface methodology. Plackett-Burman design was first used to screen the dominant impact factors for the growth of C. cellulolyticum, which were determined as yeast extract concentration, cellobiose concentration and culture temperature. The steepest ascent path design was then applied to gain the suitable range close to the optimal culture conditions for obtaining high cell density. The central composite design and the response surface analysis were finally used to determine the optimal levels of the influential factors, which were 3 g/L for yeast extract concentration, 7 g/L cellobiose concentration and 34 degrees C for culture temperature. The optimized medium was used for flask culture, and OD600 of C. cellulolyticum was increased from 0.303 to 0.586. With a pH-controlled fermentor at batch mode, OD600 reached 3.432, which was 2.8 times higher than elsewhere reported. These results support further study on the high-density culture of C. cellulolyticum and its application.
Bacteriological Techniques ; methods ; Clostridium cellulolyticum ; growth & development ; Culture Media ; Population Density

This study was aimed to analyze the results of false positive reaction in bacterial detection of blood samples with BacT/ALERT 3D system, to evaluate the specificity of this system, and to decrease the false positive reaction. Each reaction flasks in past five years were processed for bacteria isolation and identification. When the initial cultures were positive, the remaining samples and the corresponding units were recultured if still available. 11395 blood samples were detected. It is worthy of note that the incubator temperature should be stabilized, avoiding fluctuation; when the cultures were alarmed, the reaction flasks showed be kept some hours for further incubation so as to trace a sharply increasing signal to support the judgement of true bacterial growth. The results indicated that 122 samples (1.07%) wee positive at initial culture, out of them 107 samples (88.7%) were found bacterial, and 15 samples (12.3%) were found nothing. The detection curves of positive samples resulted from bacterial growth showed ascent. In conclusion, maintenance of temperature stability and avoidance of temperature fluctuation in incubator could decrease the occurrence of false-positive reaction in detection process. The reaction flasks with positive results at initial culture should be recultured, and whether existence of a sharply ascending logarilhimic growth phase in bacterial growth curve should be further detected, which are helpful to distinguish false-positive reactions from true positive, and thus increase the specificity of the BacT/ALERT system.
Automation, Laboratory ; Bacteria ; isolation & purification ; Bacteriological Techniques ; methods ; Blood ; microbiology ; Culture Media ; False Positive Reactions ; Humans

A magnetic separator was used to separate magnetic bacteria based on their magnetotactic characteristics. Acidithiobacillus ferrooxidans, a bacterium that could synthesize intra-cellular nanometer magnetic particles, was investigated as an example. Strong magnetic and weak magnetic cells were separated and collected. On average, the number of the magnetic particles present in the strong magnetic cells is more than that of the weak magnetic cells. Moreover, semisolid-plate magnetophoresis showed that the magnetotaxis of strong magnetic cells was stronger than the weak magnetic cells. These results suggest that the magnetic separator can be used to isolate the magnetic bacteria, which will facilitate the research of magnetic bacteria.
Acidithiobacillus ; isolation & purification ; metabolism ; Bacteria ; isolation & purification ; metabolism ; Bacterial Physiological Phenomena ; Bacteriological Techniques ; methods ; Magnetics

In order to develop a rapid and reliable method for B. cereus genotyping, factors influencing PFGE results, including preparation of bacterial cells embedded in agarose, lysis of embedded cells, enzymatic digestion of intact genomic DNA, and electrophoresis parameters allowing for reproducible and meaningful DNA fragment separation, were controlled. Optimal cellular growth (Luria-Bertani agar plates for 12-18 h) and lysis conditions (4 h incubation with 500 µg/mL lysozyme) produced sharp bands on the gel. Restriction enzyme NotI was chosen as the most suitable. Twenty-two isolates were analyzed by NotI digestion, using three electrophoretic parameters (EPs). The EP-a was optimal for distinguishing between isolates. The optimized protocol could be completed within 40 h which is a significant improvement over the previous methods.
Bacillus cereus ; genetics ; isolation & purification ; Bacteriological Techniques ; DNA, Bacterial ; chemistry ; genetics ; Electrophoresis, Gel, Pulsed-Field ; methods

OBJECTIVETo establish a highly efficient culture method for detecting Yersinia enterocolitica in stool samples from diarrheic patients.

METHODSStool samples collected from 200 diarrheic patients were detected with a modified and 3 conventional methods, and the positivity rates of the bacterium were compared statistically.

RESULTSWith the modified method, 18 positive samples for Yersinia enterocolitica were detected from 200 stool samples, while with the 3 conventional methods, only 3, 5, and 8 positive samples were identified, respectively. Statistical analysis with McNemer test suggested significant difference in the positive detection rate between the modified and the 3 conventional methods.

CONCLUSIONThe modified method for Yersinia enterocolitica detection has much higher sensitivity than the 3 conventional methods, and can be applied in clinical detection.

Bacteriological Techniques ; Culture Techniques ; methods ; Diarrhea ; microbiology ; Feces ; microbiology ; Humans ; Temperature ; Yersinia enterocolitica ; growth & development ; isolation & purification

Listeria monocytogenes is a pathogenic bacterium, therefore, it is essential for food safety monitoring to establish a rapid and specific detecting method. In this study, immunomagnetic beads and selective medium were combined to detect Listeria monocytogenes at different concentrations (10(1)-10(5) CFU/mL). Other three types of Listeria spp., Staphylococcus aureus and Vibrio parahaemolyticus were also detected to conduct the cross-reaction analysis. Meanwhile, contaminated milk samples were prepared to explore the limit of detection of immunomagnetic beads combining with selective medium. Results showed that Listeria monocytogenes with the concentration of 10(3) CFU/mL and above was successfully detected. Milk samples were detected within 6 hours, with a detection limit of 0.7 CFU/mL. The method developed is capable of detecting milk samples within 30 h, which is 38 h faster compared with national standard method with the same sensitivity.
Bacteriological Techniques ; methods ; Culture Media ; chemistry ; Immunomagnetic Separation ; methods ; Listeria monocytogenes ; growth & development ; isolation & purification ; Sensitivity and Specificity

BACKGROUND: Continuous monitoring systems have allowed determination of the time-to-positivity (TTP). We evaluated the clinical relevance of TTP in the BACTEC9240 system (Becton-Dickinson, USA). METHODS: A total of 2,354 vials of positive blood cultures were evaluated over 2 months. TTP was monitored from each of BACTEC Plus Aerobic/F (BD) or Pediatric Plus/F and Lytic Anaerobic/F bottles, and the differential time-to-positivity (DTP) for blood samples drawn simultaneously via catheter and a peripheral site was determined. RESULTS: The average TTP of the positive vials was 17.4 hr, and 79.9% and 95.2% of the vials showed positivity within 24 and 48 hr, respectively. While the average TTP values for Aeromonas hydrophila, Bacillus cereus, Acinetobacter baumannii, and Streptococcus pneumoniae were less than 10 hr, those for Candida spp., anaerobes, Propionibacterium acnes, Corynebacterium spp, Bacillus spp. other than cereus, and coagulase-negative staphylococci were 35.3, 27.0, 56.8, 45.8, 23.0, and 26.3 hr, respectively. The negative predictive values of TTP over 24 hr to predict Staphylococcus aureus among staphylococci and S. pneumoniae among alpha-hemolytic streptococci were 76.7% and 100%, respectively. Enterobacteriaceae and Enterococcus faecalis showed shorter TTP in anaerobic vials than in aerobic vials. DTP of more than 2 hr was observed for 27.8%, 72.2%, and 45.5% of S. aureus, S. epidermidis, and Candida spp. CONCLUSIONS: TTP can be used to discriminate pathogens and contaminants. The shorter TTP in anaerobic vials of certain Enterobacteriaceae and Enterococcus spp. would facilitate further identification. DTP is useful for diagnosing catheter-related bloodstream infection by S. aureus, S. epidermidis, and Candida spp.
Bacteremia/*diagnosis ; Bacteria, Aerobic/isolation &purification ; Bacteria, Anaerobic/isolation &purification ; Bacteriological Techniques/instrumentation/methods ; Humans ; Reagent Kits, Diagnostic ; Time Factors

BACKGROUND: We developed a new disposable anaerobic culture system, namely, the Quick anaero-system, for easy culturing of obligate anaerobes. METHODS: Our system consists of 3 components: 1) new disposable anaerobic gas pack, 2) disposable culture-envelope and sealer, and 3) reusable stainless plate rack with mesh containing 10 g of palladium catalyst pellets. To evaluate the efficiency of our system, we used 12 anaerobic bacteria. We prepared 2 sets of ten-fold serial dilutions of the 12 anaerobes, and inoculated these samples on Luria-Bertani (LB) broth and LB blood agar plate (LB-BAP) (BD Diagnostic Systems, USA). Each set was incubated in the Quick anaero-system (DAS Tech, Korea) and BBL GasPak jar with BD GasPak EZ Anaerobe Container System (BD Diagnostic Systems) at 35-37degrees C for 48 hr. The minimal inoculum size showing visible growth of 12 anaerobes when incubated in both the systems was compared. RESULTS: The minimal inoculum size showing visible growth for 2 out of the 12 anaerobes in the LB broth and 9 out of the 12 anaerobes on LB-BAP was lower for the Quick anaero-system than in the BD GasPak EZ Anaerobe Container System. The mean time (+/-SD) required to achieve absolute anaerobic conditions of the Quick anaero-system was 17 min and 56 sec (+/-3 min and 25 sec). CONCLUSIONS: The Quick anaero-system is a simple and effective method of culturing obligate anaerobes, and its performance is superior to that of the BD GasPak EZ Anaerobe Container System.
Bacteria, Anaerobic/*growth &development ; Bacteriological Techniques/instrumentation/methods ; Culture Media/chemistry ; Gases/chemistry ; Palladium/chemistry ; Reagent Kits, Diagnostic