Caffeine demethylation monitoring using a transdermal sweat patch

Biofilms consist of microorganisms immobilized at a substratum surface embedded in an organic polymer matrix of bacterial origin. Tubing drawn from the fluid pathways within dialysis machines of various models were investigated for biofilm. Scanning electron microscopy (SEM), performed on approximately 2 cm2 samples of the tubing inner surfaces revealed that the inner surfaces of the tubing were covered with biofilms consisting of numerous deposits and glycocalix at different stages of formation with components containing bacteria and algae. Evaluations of biomass were performed from tubing sections of various lengths and inner diameters put in tubes containing water for injection and immersed in an ultrasound washtub for 1 h to ensure sloughing of the biofilm. Living bacteria were identified by plating on nutrient agar media and incubation for 48 h at 37 degrees C. Epifluorescent stains were used for the total bacteria count. Lipopolysaccharide levels were determined by the endotoxin activity measurements. Polyoside contents were determined by the colometric method, and the chemical oxygen demand was measured to evaluate the amount of organic substance. Biofilms detached from tubing samples drawn from the water path, bicarbonate path, and fresh dialysate path within dialysis machines contained approximately 1.10(3)-1.10(6) total bacteria/cm2, yet only some living bacteria were found. Endotoxin levels ranged from 1 to 12 EU/cm2. In contrast in the dialysate fluid, no bacteria were found, and the endotoxin content was under the detection level of the method. The polyoside content and chemical oxygen demand of the biomass ranged from 11 to 83 microg/cm2 and from 53 to 234 mg/cm2, respectively. It is concluded that a germ- and endotoxin-free dialysate does not exclude the risks and hazards of bacteria and endotoxin discharge from biofilm developed on the fluid pathway tubing, acting as a reservoir for continuous contamination, and efforts in the optimization of cleaning and disinfection procedures used for hemodialysis systems should aim to detach and neutralize biofilm when necessary.     

POEMS syndrome in a patient with diabetic ketoacidosis: case report and review

This article summarise the main data in the literature on the role of bacteriological contamination of the dialysate fluid in inflammatory reactions in hemodialysis. Pyrogenic substances of small molecular weight from Gram-negative bacteria grown in dialysate can pass across intact dialyzer membrane to stimulate cytokine production by peripheral blood mononuclear cells. Cellulosic hemodialysis membranes are more permeable to endotoxins than synthetic membranes. Polysulfone membranes and polyamide membranes are able to adsorb bacterial toxins on the dialysate side. The diffusive transfer of bacterial products across dialysis membrane from dialysate fluid was demonstrated. Transmembrane passage of cytokine-inducing bacterial products across reprocessed dialyzers is greater than across new dialyzers. Bacteriological contamination of the dialysate fluid is a problem which must be considered with much more care by nephrologists, especially as LAL test is unable to detect all the bacterial products which can contaminate the dialysate fluid.     

Absence of bacteremia and endotoxemia despite contaminated dialyzate

Fevers associated with hemodialysis have been attributed to the transfer of relatively large endotoxin molecules and/or bacteria from contaminated dialyzate across the dialyzing membrane. We evaluated 27 patients during hollow-fiber dialysis when, due to a malfunction, dialysis fluids contained bacteria and endotoxin at levels previously reported to be associated with pyrogenic reactions. Neither endotoxin nor bacteria was detected in 54 venous and arterial blood specimens collected at the termination of hemodialysis. Temperature elevations did not occur during or within 1/2 hr after dialysis. In an extended study, 20 dialyzers were collected after single patient use and the dialyzate compartment was filled with highly contaminated dialyzate, while the blood compartment was filled with sterile pyrogen-free saline. Following 5 to 7 days incubation, bacteria were present in the blood compartments of 4 of 20 dialyzers, probably due to contamination during dialyzer handling. However, the much smaller endotoxin molecule could not be detected in the absence of bacterial contamination. These results indicate that the intact cellophane membrane is an effective barrier to endotoxin and bacteria under clinical conditions.     

Experimental investigation of the distribution of residual strains in the artery wall

Diarrheal diseases often result from ingestion of contaminated water or food. The population of La Paz, Bolivia is directly or indirectly exposed to the sewage-contaminated La Paz River. We conducted a bacteriologic survey of the La Paz River to quantify the level of bacterial contamination, with particular reference to enteropathogens. A total bacterial count exceeding 10(6) colony-forming units (CFU)/ml, including lactose fermenting and nonfermenting, gram-negative bacilli of approximately 10(5) CFU/ml, respectively, were detected in river water samples collected near two densely populated areas. A total bacterial count of 10(5) CFU/ml was also detected at the most downstream area of the river near a sparsely populated area. At four sampling locations, several enteropathogens were detected, including five enterotoxigenic Escherichia coli (ETEC) (serotype O6, O15, and O159), two enteropathogenic E. coli (EPEC) (serotype O44), two enteroinvasive E. coli (EIEC) (serotype O29), and three Salmonella O4 group isolates. The heat-labile enterotoxin gene and the invasive toxin gene were detected in all ETEC and EIEC isolates by polymerase chain reaction analysis. Nine isolates of E. coli were found by the agar dilution method to be susceptible to ampicillin, kanamycin, nalidixic acid, tetracycline, and chloramphenicol, and ampicillin resistance was found in only two isolates of EIEC 7-4 (serotype O29) and EPEC 7-5 (serotype O44). Ampicillin resistance was coded on plasmids and transferred conjugatively to E. coli chi1037 at a frequency of 10(-5) CFU/donor by the broth mating method. Strains of Aeromonas caviae, which can cause diarrheal disease in infants, were detected in vegetables grown in fields irrigated by water from the La Paz River. The survival of nine isolates of E. coli in filtered river water was compared with that of laboratory strains (E. coli chi1037, W3110, and ATCC29577). The survival time of seven isolates, excluding two ampicillin-resistant isolates, was markedly longer than that of the laboratory strains. Our results show a high bacterial contamination of the La Paz river and suggest that such levels may contribute to the high incidence of diarrheal disease in the city of La Paz.     

Removal of Indoor Airborne Bacteria by Nano-Ag/TiO2 as Photocatalyst: Feasibility Study in Museum and Nursing Institutions

Deterioration of indoor air quality attributable to airborne bacterial consortia is a widespread environmental problem. The main objective of this study is to evaluate the feasibility of applying the syngetic effect of nano-Ag/TiO2 as a photocatalyst and UV light to enhance the disinfecting capability of full-scale bacterial restraining equipment on-site in the National Museum of Natural Science and a medical-nursing institute. The influence of initial counts of total airborne bacteria and volume of space on the efficiency of bacterial restraining have been studied. In the case of museum application, a higher initial total bacterial count leads to better bacterial restraining rates; Site A (initial total bacterial counts = 506??CFU/m3) has the best bacterial restraining rate (92%) as compared with Site B (69%, initial total bacterial counts = 158??CFU/m3) and Site C (80%, initial total bacterial counts = 338??CFU/m3) after 24 h of operation. Higher initial counts of total airborne bacteria lead to an increasing bacterial restraining rate. Approximately 92% (Site A) and 74% of restrained bacterial rate were observed in a museum and nursing institutions, respectively, under the similar initial total airborne bacterial counts (506, 598??CFU/m3). The results illustrate that changes in the volume of space do not have significant inhibitory effects on the efficiency. The proposed equipment can disinfect air to restrain bacteria effectively, as demonstrated on-site in museums and nursing institutions; the results will be valuable references for designing a full-scale commercialized device for large-scale applications in the future.     

Full-scale studies of factors related to coliform regrowth in drinking water

An 18-month survey of 31 water systems in North America was conducted to determine the factors that contribute to the occurrence of coliform bacteria in drinking water. The survey included analysis of assimilable organic carbon (AOC), coliforms, disinfectant residuals, and operational parameters. Coliform bacteria were detected in 27.8% of the 2-week sampling periods and were associated with the following factors: filtration, temperature, disinfectant type and disinfectant level, AOC level, corrosion control, and operational characteristics. Four systems in the study that used unfiltered surface water accounted for 26.6% of the total number of bacterial samples collected but 64.3% (1,013 of 1,576) of the positive coliform samples. The occurrence of coliform bacteria was significantly higher when water temperatures were > 15 degrees C. For filtered systems that used free chlorine, 0.97% of 33,196 samples contained coliform bacteria, while 0.51% of 35,159 samples from chloraminated systems contained coliform bacteria. The average density of coliform bacteria was 35 times higher in free-chlorinated systems than in chloraminated water (0.60 CFU/100 ml for free-chlorinated water compared with 0.017 CFU/100 ml for chloraminated water). Systems that maintained dead-end free chlorine levels of < 0.2 mg/liter or monochloramine levels of < 0.5 mg/liter had substantially more coliform occurrences than systems that maintained higher disinfectant residuals. Free-chlorinated systems with AOC levels greater than 100 micrograms/liter had 82% more coliform-positive samples and 19 times higher coliform levels than free-chlorinated systems with average AOC levels less than 99 micrograms/liter. Systems that maintained a phosphate-based corrosion inhibitor and limited the amount of unlined cast iron pipe had fewer coliform bacteria. Several operational characteristics of the treatment process or the distribution system were also associated with increased rates of coliform occurrence. The study concludes that the occurrence of coliform bacteria within a distribution system is dependent upon a complex interaction of chemical, physical, operational, and engineering parameters. No one factor could account for all of the coliform occurrences, and one must consider all of the parameters described above in devising a solution to the regrowth problem.     

The use of heated citric acid for dialyzer reprocessing

Dialyzer reprocessing with heated water (100 to 105 degrees C) for 20 h can be used safely in lieu of chemical methods for disinfection. All infective agents including spores are destroyed and depyrogenation may occur. However, these temperatures may result in structural damage to the dialyzer, limiting reuse. Dialyzer reprocessing by using 1.5% citric acid heated to 95 degrees C for 20 h is an alternative method that produces equivalent microbiologic effects. Citric acid is well known as a disinfecting agent used for dialysis equipment. Because there is little structural damage to dialyzer components at 95 degrees C, reuse statistics are improved (mean reuse increased to 12.8). Both small and large molecule clearances and the sieving coefficient for protein are insignificantly altered by the process. Whereas the procedure is relatively simple, quality-assurance indicators are essential. The method has appeal because it avoids the use of chemical germicides. However, at present it has only been tested thoroughly in polysulfone dialyzers with heat-resistant polycarbonate casings and polyurethane resin. The clinical experience is favorable.     

Effects of lactobacilli on yeast-catalyzed ethanol fermentations

Normal-gravity (22 to 24 degrees Plato) wheat mashes were inoculated with five industrially important strains of lactobacilli at approximately 10(5), approximately 10(6), approximately 10(7), approximately 10(8), and approximately 10(9) CFU/ml in order to study the effects of the lactobacilli on yeast growth and ethanol productivity. Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus #3, Lactobacillus rhamnosus, and Lactobacillus fermentum were used. Controls with yeast cells but no bacterial inoculation and additional treatments with bacteria alone inoculated at approximately 10(7) CFU/ml of mash were included. Decreased ethanol yields were due to the diversion of carbohydrates for bacterial growth and the production of lactic acid. As higher numbers of the bacteria were produced (depending on the strain), 1 to 1.5% (wt/vol) lactic acid resulted in the case of homofermentative organisms. L. fermentum, a heterofermentative organism, produced only 0.5% (wt/vol) lactic acid. When L. plantarum, L. rhamnosus, and L. fermentum were inoculated at approximately 10(6) CFU/ml, an approximately 2% decrease in the final ethanol concentration was observed. Smaller initial numbers (only 10(5) CFU/ml) of L. paracasei or Lactobacillus #3 were sufficient to cause more than 2% decreases in the final ethanol concentrations measured compared to the control. Such effects after an inoculation of only 10(5) CFU/ml may have been due to the higher tolerance to ethanol of the latter two bacteria, to the more rapid adaptation (shorter lag phase) of these two industrial organisms to fermentation conditions, and/or to their more rapid growth and metabolism. When up to 10(9) CFU of bacteria/ml was present in mash, approximately 3.8 to 7.6% reductions in ethanol concentration occurred depending on the strain. Production of lactic acid and a suspected competition with yeast cells for essential growth factors in the fermenting medium were the major reasons for reductions in yeast growth and final ethanol yield when lactic acid bacteria were present.     

The synaptic process in Locusta migratoria spermatocytes by synaptonemal complex analysis

BACKGROUND: Bacterial contamination of treated water and dialysate comprises an important problem for patients undergoing haemodialysis. Both the progressive reduction of the thickness of cellulose membranes and the expanding use of high-flux membranes probably enhance the risk of pyrogenic reactions, therefore increasing the need for atoxic water and non-pyrogenic dialysis fluid. METHODS: Samples of tap water, treated water, and effluent dialysate in all 85 haemodialysis centres in Greece were examined for total heterotrophic bacteria counts employing the pour plate method, total and faecal coliforms, faecal streptococci and pseudomonas spp. using the membrane filter technique, and sulphite-reducing clostridia applying the most probable number method. Overall 255 paired samples were tested from January to March 1997. RESULTS: For total heterotrophic bacteria, the overall compliance of treated water and dialysate to the American Association of Medical Instrumentation standards (<200 c.f.u./ml for water and <2000 c.f.u./ml for dialysate) was 92.6 and 63.7% respectively, whereas the compliance of tap water samples to our national standards (total heterotrophic bacteria < 10 c.f.u./ml and absence of the other indicator bacteria) was 80.7%. The most commonly isolated bacteria were pseudomonas spp., found in 22.2% of treated water and 59.5% of dialysate samples, whereas the respective frequencies were 12.3 and 36.2% for total coliforms, 8.6 and 30.0% for faecal coliforms, 14.8 and 28.7% for faecal streptococci, and sulphite-reducing clostridia were isolated in 5.8% of dialysate samples only. Haemodialysis centres equipped with storage tanks for treated water experienced lower levels of total heterotrophic bacteria, but higher counts of total and faecal coliforms, faecal streptococci, and pseudomonas spp., although the difference was statistically significant only for faecal streptococci counts, (P<0.05). Sixty-seven haemodialysis centres were equipped with bacterial filters, but mean values of all the examined microorganisms were not statistically different from those of the other centres. Faecal streptococci counts in treated water samples were positively correlated with ageing of both haemodialysis centres (P<0.005) and purification system (P<0.05), whereas pseudomonas counts were significantly correlated with ageing of the purification system (P<0.05).     

Air quality and microbiologic contamination in operating theatres

The present study concerns the air quality and microbiological contamination in two newly built operating theatres; one with laminar air flow (LAF) equipment for cardio-thoracic operations, and one with conventional ventilation for urological operations. Both theatres had an identical number of air exchanges (17/h), identical microclimatic conditions and they employed the same cleaning procedures. In the LAF-ventilated operating theatre bacterial contamination of the air was effectively reduced to less than 10 colony-forming units (CFU)/m3 in all 125 samples (1 m3 per sample) tested. In most samples, 118/125, the bacterial count was less than 5 CFU/m3, despite the presence of ten persons. The conventionally ventilated theatre reached values up to 120 CFU/m3 during the most active period of the day when approximately seven persons were present. The LAF ventilation reduced both the content of particles in the air and contamination by bacteria on the floor. In both theatres cleaning procedures had only a low impact on CFU in the air and on the floor. The use of diathermia markedly increased the level of small particles in the air, and this may influence the air quality in the operating theatres.     

Magnetic resonance tomography in neurocutaneous melanosis

Beef lean, fat, and connective tissues were inoculated with Escherichia coli O157:H7 before and after a prewashing procedure to compare the efficacy of prewashing and no prewashing on bacterial adherence and, consequently, on the removal of bacteria from the inoculated surfaces. Prewashing consisted of spraying tissues with tap water before inoculation. Final washing with disinfectant solutions compared the efficacy of several chemicals for the removal or destruction of E. coli O157:H7. The results showed that prewashing was very effective in reducing the numbers of bacterial cells on beef tissues, mainly lean tissue, in the control samples which received final washing with water. An opposite effect of prewashing was observed when disinfectant solutions were used for final washing; this may be due to dilution by water carried on the tissues after prewashing. The efficacy of chemicals was dependent on the type of exposed tissue. Hydrogen peroxide (3%) was more efficient in the removal of E. coli O157:H7 from connective tissues, with reductions greater than 4 log CFU/cm2, compared to a normally washed control (P < 0.01). Chlorhexidine (0.1%) was very efficient on fat and lean tissues, causing reductions over 5 log CFU/cm2 on not prewashed fat and lean tissues, compared to the control (P < 0.01). Acetic acid (5%) was the least effective, decreasing the number of CFU by under 1 log/cm2 as compared to the control; and no statistically significant difference was found among tissues, even though the removal of bacteria seemed less in lean tissue compared to fat or connective tissues.     

Transfusion-related bacterial sepsis

Transfusion-related bacterial sepsis, although infrequent, is a serious and sometimes fatal transfusion complication. Several new studies confirm previous observations of the prevalence of contamination in red cell and platelet components. In addition, several recent reports have described the risk of bacterial contamination of hematopoietic progenitor preparations. Other studies have investigated potential interventions including development of alternative skin cleansing methods, alteration of whole blood storage time, cold storage of platelet suspensions, leukoreduction of red cells, development of rapid screening tests for bacteria, and a method for bacterial inactivation of platelet components. A generic approach to inactivation, such as those targeting bacterial nucleic acid, is particularly attractive because this method would not depend on the strain or source of contamination.     

Convective treatments with on-line production of replacement fluid: a clinical experience lasting 6 years

BACKGROUND: The introduction of techniques with on-line (OL) production of replacement fluid by filtration of dialysis fluid raises concerns about exposure of dialysis patients to pyrogenic substances. This work was undertaken to evaluate safety and feasibility of OL preparation of replacement fluid for haemodiafiltration (HDF). METHODS: OL HDF was carried out with commercially available monitors without any adjustment in the operational organization of our Centre. Bicarbonate dialysis fluid was filtered twice before being reinjected into the patients. The effects of acute load of OL fluid were assessed by very sensitive in vitro and in vivo tests; the chronic effects were assessed by monitoring the patients for the appearance of any untoward clinical manifestations and by measuring their cytokine response. RESULTS: In a pilot study the membrane filter culture technique of replacement fluid yielded no bacteria or mycetes growth, while LAL test was < 0.01 EU/ml. The normal human monocyte production of TNF alpha, IL-1 beta and IL-1Ra was not significantly different when cells were incubated with OL or commercial replacement fluid. The patients' body temperature profile (continuous recording during treatments and the following 24 h) overlapped with that of the control procedure. Over 6 years we performed 4284 OL treatments (total amount reinjected fluid 102,900 litres) on 13 patients treated for 26 +/- 9 months. In none of these treatments did we observe pyrogenic reactions. In comparison with the previous period on standard bicarbonate haemodialysis, OL HDF afforded significantly better cardiovascular tolerance to fluid removal and higher Kt/V values. The nutritional status did not deteriorate, while the acute-phase reactants and serum beta 2M levels did not increase. Moreover, no translucent cysts or destructive arthropathy were observed on bone X-rays. The patients' plasma cytokine levels and monocytes cytokines production, measured either before or after a single OL HDF, were comparable with the values obtained in controls treated with standard HDF. CONCLUSIONS: We conclude that OL-prepared replacement fluid is as safe as that of the commercial bags with regard to sterility and non-pyrogenicity. OL HDF can be readily implemented in any dialysis centre without bringing any further burden on the staff.     

Bacterial tracking in a dairy production system using phenotypic and ribotyping methods

A systematic sampling plan was designed to collect raw and pasteurized milk samples throughout a single-raw milk source, dairy-processing operation experiencing reduced product shelf lives due to bacterial contamination. The objectives were to track bacterial contamination sources throughout a complete dairy production system and use this information to reduce bacterial spoilage losses in processed fluid products. Over a 5-week period, 233 bacterial isolates were collected, representative of different colony morphologies on psychrotrophic bacteria count (PBC) plates. Forty-five isolates (19%) were obtained from pasteurized milk and 188 (81%) were isolated from raw product. Thirty isolates were identified as Pseudomonas spp. by Gram stain and biochemical methods. Of these, 27 (90%) were postpasteurization isolates and 3 (10%) were raw milk isolates. Automated ribotyping revealed that raw and pasteurized Pseudomonas fluorescens isolates were indistinguishable (similarity index > 0.93), suggesting the possibility of postpasteurization contamination with bacteria from raw product. In the plant, filler nozzles were identified as the primary reservoirs of postpasteurization contamination. Nozzle replacement produced significantly lower finished-product PBCs at 7 days postprocessing (> 4-log reduction) and extended fluid product shelf life.     

Automated peritoneal dialysis with 'on-line'-prepared bicarbonate-buffered dialysate: technique and first clinical experiences

BACKGROUND: Automated peritoneal dialysis (APD) has the possibility of increasing the dialysis efficacy by using higher fill volumes, frequent dialysate exchanges, and tidal techniques. It is then possible to treat patients adequately without residual renal function. The drawbacks of the required high amounts of dialysis solution of up to 30 litres per session are the high costs of lactate-based dialysate bags and difficulties for the patients in handling these bags. So far, bicarbonate-based peritoneal dialysate, which may be more biocompatible, is only available for CAPD in double-chamber bags. In APD this could be overcome by 'on-line' preparation of bicarbonate-buffered dialysate using advanced technologies originally designed for on-line preparation of substitution fluid for haemofiltration. METHODS: Four patients without residual renal function were treated with APD five times weekly in a crossover study design. Patients received standard lactate-based (35 mmol/l) treatment (25 litres per session each) in weeks 1 and 3. In week 2 on-line-produced bicarbonate-buffered (37 mmol/l) dialysate was used. This dialysate was prepared by an AK 100 Ultra haemodialysis machine. The machine was modified for adding glucose from a 50% concentrate to the desired concentration of 1.7%. Electrolytes, pH, pCO2, and dialysis efficacy parameters were measured. Microbiological testing was carefully performed. RESULTS: Creatinine clearances, Kt/V, and pCO2 did not vary between the different treatment phases, whereas the pH showed a distinct increase during the bicarbonate phase. Repeated determinations of endotoxins and culturing showed no contamination of the dialysate. The composition of the produced dialysate was reproducible with respect to pH, pCO2, sodium, calcium and bicarbonate, whereas the glucose concentration varied by +/- 20%. CONCLUSIONS: On-line preparation of PD fluid with the AK 100 Ultra is easy and safe to handle. APD with dialysate containing 37 mmol/l bicarbonate provides improved acid base balance and possibly improved biocompatibility, and may lead to a significant cost reduction. Further development in order to provide smaller machines and more precise ways of achieving a desired dialysate glucose concentration is necessary.     

Culture of dialysis fluids on nutrient-rich media for short periods at elevated temperatures underestimate microbial contamination

Recommended culture methods for monitoring bacterial contamination of H2O, dialysate and bicarbonate concentrate in dialysis centers in the USA involves culturing these fluids for 48 h at 37 degrees C. A variety of media and commercial culture methods are accepted for monitoring these fluids. Over a 3-month a comparison was made between an acceptable culture method, tryptic soy agar (TSA) employing the pour plate (PP) technique at 37 degrees C for 48 h, and PP cultures on standard methods agar (SMA) and R2A agar, incubated at ambient temperature (23 degrees C) for 48, 72, 168 h. Increases in the colony counts over time occurred for all three fluids. However, counts wee greater on SMA and R2A than on TSA. The increases over the standard 48-hour TSA cultures ranged as high as 10(4) times for 23 degrees C cultures at 7 days of incubation. Endotoxin levels even in the most contaminated samples were found to be below the acceptable 5 EU/ml recommended for reprocessor water. Bacterial colonies that appeared at 48, 72 and 168 h were isolated and identified. Pseudomonas, Moraxella, Acinetobacter and CDC group VI C-2 were among some of the common bacteria isolated. This study indicates that the media utilized, the time and temperature of incubation may result in a significant underestimation of the bacterial population of water and dialysis fluids, thus potentially placing the patient at a higher risk.     

Distribution and Transport of Fecal Bacteria at Spring Thaw in a Rural Alaskan Community

People living without piped water and sewer can be at increased risk for diseases transmitted by the fecal-oral route. One Alaskan community that relies on hauled water and sewage was studied to determine the pathways of fecal contamination of drinking water and the human environment so that barriers can be established to protect human health. Samples were tested for the fecal indicators Escherichia coli and Enterococcus. Several samples were tested for the pathogens Giardia lamblia and Cryptosporidium parvum and source tracking methods were employed. Surface water flow transported bacteria within the community during spring thaw and human fecal contamination was detected in town, but flow from the dump did not appear to contribute to contamination in town. Within the home, fecal bacteria were found on water dippers, kitchen counters and floors, and in hand-washing basins. Giardia was found at the dump, but not in water from the river adjacent to the community. Exposure to fecal contamination could be reduced by cleaning up after dogs, carefully disposing of wastewater, and by protecting stored drinking water.     

Quantitation of bacteria in blood of typhoid fever patients and relationship between counts and clinical features, transmissibility, and antibiotic resistance

Salmonella typhi was isolated from 369 and Salmonella paratyphi A was isolated from 6 of 515 Vietnamese patients with suspected enteric fever. Compared with conventional broth culture of blood, direct plating of the buffy coat had a diagnostic sensitivity of 99.5% (95% confidence interval [CI], 97.1 to 100%). Blood bacterial counts were estimated by the pour plate method. The median S. typhi count in blood was 1 CFU/ml (range, <0.3 to 387 CFU/ml), of which a mean of 63% (95% CI, 58 to 67%) were intracellular. The mean number of bacteria per infected leukocyte was 1.3 (interquartile range [IQR], 0.7 to 2.4) CFU/cell (n = 81). Children (< 15 years old; n = 115) had higher median blood bacterial counts than adults (n = 262): 1.5 (range, <0.3 to 387) versus 0.6 (range, <0.3 to 17.7) CFU/ml (P = 0.008), and patients who excreted S. typhi in feces had higher bacteremias than those who did not: a median of 3 (range, <0.3 to 32) versus 1 (range, <0.3 to 68) CFU/ml (P = 0.02). Blood bacterial counts declined with increasing duration of illness (P = 0.002) and were higher in infections caused by multidrug-resistant S. typhi (1.3 [range, <0.3 to 387] CFU/ml; n = 313) than in infections caused by antibiotic-sensitive S. typhi (0.5 [range, <0.3 to 32] CFU/ml; n = 62) (P = 0.006). In a multivariate analysis this proved to be an independent association, suggesting a relationship between antibiotic resistance and virulence in S. typhi.     

Host defences in continuous ambulatory peritoneal dialysis and the genesis of peritonitis

Continuous ambulatory peritoneal dialysis (CAPD) has come to be extensively used for the treatment of end-stage renal failure in children, and especially infants, such that now more than half of children on dialysis worldwide receive treatment by this means. Peritonitis, however, is commoner in children than in adults receiving treatment, and is a major source of morbidity and treatment failure in children started on CAPD. Only recently has the immunology of the normal peritoneum been studied extensively, with the need to assess the impact of the installation of large volumes of fluid into the peritoneal sac during dialysis. The main phagocytic defences of the peritoneum depend upon a unique set of macrophages which are present free in the peritoneal fluid but also in the submesothelium and in perivascular collections together with B lymphocytes in the submesothelial area. Both the number of macrophages per unit volume and the concentration of opsonic proteins, such as IgG, complement and fibronectin, are reduced to between only 1% and 5% when dialysis fluid is continuously present in the peritoneal sac. In addition, the fluids used for CAPD are toxic to both macrophages and to mesothelial cells. Thus minor degrees of contamination frequently lead to peritonitis and in addition the majority of patients have catheters inserted in their peritoneum which become colonised with organisms capable of producing exopolysaccharide (slime), which promotes adhesion of the organism to the plastic and protects them against phagocytic attack and the penetration of antibiotics. Thus the peritoneum is in a state of continual inflammation, as well as being a markedly more vulnerable site than the normal peritoneum to the entry of organisms. Whether clinical peritonitis appears in this state of chronic contamination probably depends on perturbation in the balance between host defences and the organism. Whilst Staphylococcus epidermidis is the commonest cause of peritonitis, Staphylococcus aureus and Gram-negative organisms are much more serious and more frequently lead either to temporary catheter removal or discontinuation of dialysis altogether. This review describes the peritoneal defences in relation to the genesis of peritonitis.     

Transmembrane passage of cytokine-inducing bacterial products across new and reprocessed polysulfone dialyzers

The widespread use of bicarbonate dialysate and reprocessed high-efficiency and "high-flux" dialyzers has raised concerns about the increased risk of reverse-transfer of dialysate contaminants into the blood compartment. To evaluate this concern, the reverse-transfer of bacterial products from contaminated bicarbonate dialysate into the blood compartment was compared during in vitro dialysis with new or reprocessed high-flux polysulfone dialyzers. In vitro dialysis was carried out at 37 degrees C by use of a counter-current recirculating loop dialysis circuit with either new high-flux polysulfone dialyzers or dialyzers reprocessed once or 20 times with formaldehyde (0.75%) and bleach (< 1%) with an automated system. Heparinized whole blood from healthy volunteers was circulated through the blood compartment, and bicarbonate dialysate was circulated in the dialysate compartment. The dialysate was challenged sequentially by 1:1000 and 1:100 dilutions of a sterile Pseudomonas aeruginosa culture supernatant (bacterial challenge). Samples were drawn from the blood and dialysate compartments 1 h after each challenge. Peripheral blood mononuclear cells (PBMC) were harvested by Ficoll-Hypaque separation from whole blood in the blood compartment and a 5 x 10(6) PBMC/mL cell suspension was prepared. Likewise, dialysate samples (0.5 mL) were added to 0.5 mL suspension of 5 x 10(6) PBMC/mL drawn at baseline. All PBMC suspensions were incubated upright in a humidified atmosphere at 37 degrees C with 5% CO2 for 24 h, and total interleukin-1 alpha (IL-1 alpha) and tumor necrosis factor-alpha (TNF alpha) cytokine production (cell-associated and secreted) was measured by radioimmunoassay. Eight experiments were performed for each arm of the study with the same donor for each arm. One hour after contaminating the dialysate with a 1:1000 dilution of the bacterial challenge, IL-1 alpha production by PBMC harvested from the blood compartment was 160 +/- 0, 171 +/- 11, and 270 +/- 35 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.004). One hour after challenging the dialysate with 1:100 dilution, IL-1 alpha production by PBMC harvested from the blood compartment was 188 +/- 20, 228 +/- 35, and 427 +/- 67 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.006). IL-1 alpha production by PBMC from dialyzers reprocessed 20 times was significantly greater than both new and dialyzers reprocessed once. However, there were no significant differences between new dialyzers and dialyzers reprocessed once. Similarly, after the 1:1000 challenge, TNF alpha production by PBMC harvested from the blood compartment was 160 +/- 0, 160 +/- 0, and 213 +/- 22 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.008). After the 1:100 challenge, TNF alpha production was 168 +/- 8, 188 +/- 20, and 225 +/- 32 pg, respectively, for new dialyzers, dialyzers reprocessed once, and dialyzers reprocessed 20 times (P = 0.20). These results demonstrate that reprocessing of high-flux polysulfone dialyzers with bleach increases the risk of reverse-transfer of bacterial products from contaminated dialysate, and this risk appears to increase with the number of reuses. Consequently, units that reprocess membranes with bleach and have suboptimal water quality might subject their patients to a higher risk of cytokine-related morbidity.