Evaluation of Smear Layer Removal and Antimicrobial Efficacy of HybenX Against Enterococcus Faecalis Biofilm.

To evaluate the antibacterial efficacy of HybenX, an endodontic irrigant against Enterococcus faecalis biofilm and determine its efficacy in removal of smear layer. Forty root canals were inoculated with E. faecalis for three weeks, divided into 4 groups and irrigated with: Group1: 5 ml 2.5% sodium hypochlorite; Group2: 5 ml 2% chlorhexidine gluconate; Group3: 5 ml HybenX; Group4: 5 ml distilled water (control). Dentin shavings were collected and plated on agar plates, followed by colony forming unit (CFU) determination. For smear layer removal examination, 30 single rooted teeth were instrumented and divided into 3 groups and treated with: GroupI: 5 ml of 17% EDTA; GroupII:5 ml of HybenX; and GroupIII: 5 ml of distilled water. Samples were then subjected to SEM examination. All irrigants showed a significant reduction in CFUs compared to the control group (P⟨ .001), but none compared to each other. Regarding smear layer removal in the apical third, EDTA removed smear layer more effectively than HybenX (P = .014). HybenX demonstrated good antimicrobial efficacy against E. faecalis biofilm and removed the smear layer effectively in coronal and middle third of the root canal system. HybenX can be considered as a promising irrigating agent in root canal treatment of infected teeth.

Epididymal and prostatic histoplasmosis in a renal transplant recipient from southern India.

An unusual case of epididymal and prostatic histoplasmosis is reported from Karnataka State of southern India, in a 37-year-old male renal transplant recipient, who had presented with gastrointestinal histoplasmosis a year before the second presentation and had responded successfully to 9 months' treatment with itraconazole. The patient presented a second time with high-grade fever and pain in the scrotum and the lower abdomen. Ultrasound of the abdomen revealed a retrotrigonal abscess, which was confirmed on computed tomography scan of the pelvis. Pus was drained via transrectal route and biopsy specimens were taken from the epididymis and the prostate. Histopathology revealed the picture suggestive of histoplasmosis, and the pus on culture confirmed the infection with Histoplasma capsulatum. To our knowledge, only 5 cases of epididymal histoplasmosis have been reported in the literature, our case being the second report from India and sixth in the world; prostatic histoplasmosis is also a rare occurrence.

Microbial ingress through breaches in aseptic manufacturing systems: experimental investigation of pressure-driven leaks of liquids.

The ability of pressure-driven leaks to provide an effective barrier against microbial ingress was investigated experimentally. The leak was simulated by circulating growth promoting media through a section of flexible tubing that had a 3/4-inch breach. The flow rate of the media was adjusted until the leak was barely visible; the leak rate was minimized to enhance the possibility of microbial ingress through the breach. The site of the breach was repeatedly inoculated with an actively growing population of Pseudomonas aeruginosa--a highly motile bacterium--suspended in a nutrient-rich medium. The breach was exposed to approximately 140 million organisms over a period of 8 h. This level of contamination greatly exceeds the microbial exposure that a breach would be subjected to in an aseptic manufacturing environment. The data show that microbial ingress did not occur under the above conditions. This indicates that pressure-driven leaks in flexible tubing can provide an effective barrier against microbial ingress, and that the integrity of the sterile boundary can be maintained during such leaks. The results of this investigation could be used to perform risk assessment analyses of the potential for microbial ingress through breaches in aseptic manufacturing systems.

Effect of dissolved oxygen control on growth and antibiotic production in Streptomyces clavuligerus fermentations.

A proportional-integral control system was used to control dissolved oxygen in a fermentor at constant shear and mass transfer conditions. Growth and antibiotic production in Streptomyces clavuligerus were studied at different dissolved oxygen levels during the fermentation. Three protocols were employed: no-oxygen control to provide a base case, oxygen controlled to a preset saturation level throughout the fermentation, and oxygen controlled at a high level only during the growth phase. The last protocol was aimed at optimizing the consumption of oxygen. Lower specific growth rates and cephamycin C yields were obtained when dissolved oxygen was controlled at 50% throughout the fermentation, compared to the base case. A 2.4-fold increase in the final cephamycin yield was observed when dissolved oxygen was controlled at saturation levels during the growth phase, compared to the experiments without dissolved oxygen control. This enhancement in yield was independent of the dissolved oxygen (DO) level after exponential growth, in the range of 50-100% saturation. The most effective control strategy, therefore, was to control DO only during active growth when the biosynthetic enzymes were probably synthesized.

Experimental simulation of dissolved oxygen fluctuations in large fermentors: effect on Streptomyces clavuligerus.

An experimental Monte Carlo method was used to study the effect of fluctuations in oxygen concentration on the synthesis of antibiotics by Streptomyces clavuligerus. Air was supplied to the culture in a 2-L fermentor in random cycles following the lognormal distribution in order to model the circulation within large production-scale vessels. Each cycle consisted of air supply for 5 s followed by no aeration for the balance of the cycle time which ranged from 8 to 44 s, with a mean time of 20 s. Comparable experiments were also conducted with constant period cycling of air and with continuous supply of air. The yields of cephamycin C and its precursor, penicillin N, were suppressed by the Monte Carlo simulation of circulation in a large tank, as compared to constant period cycling. The concentration of dissolved oxygen remained at a low, ca. 5% of saturation, for 5-10 h longer during the Monte Carlo experiment than during the periodic aeration experiment. The biosynthetic enzymes, which are sensitive to oxygen levels, were likely affected not only by the mean time of cycling but also by the distribution of the cycles.

Kinetics of CO2 hydration in fermentors: pH and pressure effects.

Fluctuations in pH and head-space pressure in a fermentor introduce temporary changes in off-gas CO2 concentrations. These changes are quantified using a simple model based on kinetics of CO2 hydration and gas-liquid mass transfer. The model is verified experimentally. An eigenvalue analysis of the model indicates that mass transfer is the parameter which controls the dynamics of CO2 equilibration.

Selection of bioprocess simulation software for industrial applications.

Two commercially available, process-simulation software packages (Aspen Batch Plus v1.2, Aspen Technology, Inc., Cambridge, Massachusetts, and Intelligen SuperPro v3.0, INTELLIGEN, INC., Scotch Plains, Ner Jersey) are evaluated for use in modeling industrial, biotechnology processes. Software is quantitatively evaluated by Kepner-Tregoe Decision Analysis (Kepner and Tregoe, 1981). This evaluation shows that Aspen Batch Plus v1.2 (ABP) and Intelligen SuperPro v3.0 (ISP) can successfully perform specific simulation tasks but do not provide a complete model of all phenomena occurring within a biotechnology process. Software is best suited to provide a format for process management, using material and energy balances to answer scheduling questions, explore equipment change-outs, and calculate cost data. The ability of simulation software to accurately predict unit operation scale-up and optimize bioprocesses is limited. To realistically evaluate the software, a vaccine manufacturing process under development at Merck & Company is simulated. Case studies from the vaccine process are presented as examples of how ABP and ISP can be used to shed light on real-world processing issues.