A method for assaying biofilm capacity on polyurethane-coated slides.

Isolates of coagulase-negative Staphylococci (CNS) were examined for their ability to form biofilms on polyurethane-coated slides. These slides provided a smooth plastic coating simulating polymeric plastic surfaces of medical grade catheter tubing. Slides were placed into plastic conical tubes containing tryptic soy broth inoculated with 10(6) bacteria per mL. The tubes were then incubated at 37 degrees for 48 hours. After incubation, 1 of the slides was stained with a fluorescent acridine orange stain and the other with a safranin stain. The incubation tubes were also stained with safranin. Forty-eight percent of the 65 CNS isolates were found to form a biofilm using acridine orange staining. Forty percent of the 65 CNS isolates were found to form a biofilm using the safranin stain on slides, whereas only 34% were found to adhere on sides of plastic tubes. Increased sensitivity of the fluorescent stain was probably due to enhanced visualization of smaller numbers of bacteria on the plastic. This method using fluorescent stained plastic-coated slides was easier to visualize and interpret than the tube method.

Effectiveness of microwave moist-heat intraluminal disinfection of CAPD connectology.

Studies were performed to quantify the microbial population reduction achieved with the Peritoneal Dialysis Moist Heat Device (PDM-1). This microwave method is used to disinfect the inner lumen of connectors used in the exchange process for continuous ambulatory peritoneal dialysis (CAPD). The microwave heating technique was evaluated on different connector systems containing a suspension of 10(6) microorganisms. The most prevalent and most problematic peritonitis-causative microorganisms were tested. After heating, the degree of disinfection was measured by enumerating bacterial colonies of the treated suspension. D-value determinations were then performed. The D-value for spores of A. niger was found to be 7.1 seconds for one type of connector system. Two other connector systems containing different intraluminal solution volumes were also tested using spores of A. niger and D-values were determined to be 7.6 seconds and 9.5 seconds, respectively. Other organisms tested were determined to have D-value times shorter than that for A. niger. Rapid heating of the solution contained within the CAPD connectors is a key characteristic of the microwave technique since temperatures rise high enough for destruction of microorganisms while leaving the plastic safe to touch. Thus, this technique is a safe and effective method for providing intraluminal disinfection.

Moist heat intraluminal disinfection of CAPD connectors.

A moist heat technique for disinfecting the inner lumen of commercially available connectology used in the exchange process for Continuous Ambulatory Peritoneal Dialysis (CAPD) was evaluated. Moist heat was generated by a device (PDM-1) that directed microwave energy to heat a sample solution containing a concentration of 10(6) microorganisms inside a pair of mated plastic CAPD connectors. Microorganisms tested included those most prevalent and most problematic in causing peritonitis. Testing, performed according to F.D.A. approved standards, involved heating a sample solution and then placing the sample solution into vials which were then sealed and incubated. Absolute determination of growth versus no growth was measured by macroscopic observation. Positive control samples were performed in the same manner but were not exposed to heat. Negative controls were performed in the same manner in the absence of test organisms. At temperatures of approximately 100 degrees C a D-value of 6.6 seconds was determined using the organism found the most thermoresistant. A cycle time of 54 seconds appeared sufficient to achieve a 10(6) population reduction of all microorganisms tested. The moist heat technique offers a safe, effective method for disinfection of the inner lumen of CAPD connectors.

Studies on interference with tolerance induction in T cells.

The mechanism of T-cell tolerance to a thymus-dependent antigen was examined, using the adjuvant polyadenylic-polyuridylic acid (poly A:U). In adoptive transfer experiments, thymus cells obtained from donor mice 2 days after treatment with a tolerogenic dose of bovine gamma-globulin (sBGG) did not cooperate with bone marrow (BM) cells in irradiated recipients challenged with aggregated BGG (aBGG). In contrast, thymus cells from donors given sBGG plus poly A:U retained their helper activity, as assayed by hemagglutination and rosette formation of spleen cells. The effect of poly A:U in preventing tolerance induction was also demonstrable in the cortisone-resistant population, in that thymus cells from cortisone-treated donors that had received sBGG and poly A:U retained their helper function. The presence of suppressor cells and the effect of poly A:U on their stimulation were also examined. sBGG-treated thymus cells suppressed the response of BGG-primed spleen cells in lethally irradiated mice, whereas thymus cells from donors treated with sBGG and poly A:U were not suppressive. These observations show that poly A:U prevents tolerance induction and the development of suppressor activity in T cells.