Comparison of viral isolation and multiplex real-time reverse transcription-PCR for confirmation of respiratory syncytial virus and influenza virus detection by antigen immunoassays.

We evaluated the Prodesse ProFlu-1 real-time reverse transcription-PCR multiplex assay with the SmartCycler instrument for the detection of human respiratory syncytial virus (RSV) and influenza A and B viruses in comparison to conventional cell culture and antigen immunoassays with the BD Directigen A+B and Binax NOW RSV assays over two successive respiratory virus seasons. Ninety-two percent of the 361 specimens tested were nasopharyngeal aspirates obtained from individual patients, of which 119 were positive for RSV and 59 were positive for influenza virus. The median age of the patients whose specimens were positive for RSV and influenza virus were 6.3 months and 42.4 years, respectively. The specificity of all of the methods tested was >or=99%, and the individual sensitivities of NOW RSV, RSV culture, Directigen A+B, influenza virus culture, and the Proflu-1 PCR for influenza/RSV were 82% (95% confidence interval [CI], 73 to 88), 57% (95% CI, 44 to 69), 59% (95% CI, 44 to 72), 54% (95% CI, 38 to 69), and 98% (95% CI, 93 to 100)/95% (95% CI, 85 to 99), respectively. In a clinical setting where viral isolation is performed to confirm rapid antigen immunoassay results for these common respiratory viruses, one-step real-time reverse transcriptase PCR testing can be a more sensitive and timely confirmatory method.

Factors associated with critical-care healthcare workers' adherence to recommended barrier precautions during the Toronto severe acute respiratory syndrome outbreak.

OBJECTIVE: To assess factors associated with adherence to recommended barrier precautions among healthcare workers (HCWs) providing care to critically ill patients with severe acute respiratory syndrome (SARS). SETTING: Fifteen acute care hospitals in Ontario, Canada. DESIGN: Retrospective cohort study. PATIENTS: All patients with SARS who required intubation during the Toronto SARS outbreak in 2003. PARTICIPANTS: HCWs who provided care to or entered the room of a SARS patient during the period from 24 hours before intubation until 4 hours after intubation. METHODS: Standardized interviews were conducted with eligible HCWs to assess their interactions with the SARS patient, their use of barrier precautions, their practices for removing personal protective equipment, and the infection control training they received. RESULTS: Of 879 eligible HCWs, 795 (90%) participated. In multivariate analysis, the following predictors of consistent adherence to recommended barrier precautions were identified: recognition of the patient as a SARS case (odds ratio [OR], 2.5 [95% confidence interval {CI}, 1.5-4.5); recent infection control training (OR for interactive training, 2.7 [95% CI, 1.7-4.4]; OR for passive training, 1.7 [95% CI, 1.0-3.0]), and working in a SARS unit (OR, 4.0 [95% CI, 1.8-8.9]) or intensive care unit (OR, 4.3 [95% CI, 2.0-9.0]). Two factors were associated with significantly lower rates of consistent adherence: the provision of care for patients with higher Acute Physiology and Chronic Health Evaluation (APACHE) II scores (OR for score APACHE II of 20 or greater, 0.4 [95% CI, 0.28-0.68]) and work on shifts that required more frequent room entry (OR for 6 or more entries per shift, 0.5 [95% CI, 0.32-0.86]). CONCLUSIONS: There were significant deficits in knowledge about self-protection that were partially corrected by education programs during the SARS outbreak. HCWs' adherence to self-protection guidelines was most closely associated with whether they provided care to patients who had received a definite diagnosis of SARS.

Prevalence of nosocomial infections in high-risk units in the university clinical center of Kosova.

A point-prevalence study was performed to determine the prevalence of nosocomial infections in University Clinical Center of Kosova. Of 167 surveyed patients, 27 had a total of 29 nosocomial infections, with an overall prevalence rate of 17.4%. Nosocomial bloodstream infections were most prevalent (62%). The prevalence was highest among neonates in intensive-care units (77.8%).

Infection control in Kosova.

This article presents the authors' views on infection control in hospitals in Kosova, focusing on the main problems and appropriate solutions. Past and present government initiatives are briefly presented, and available data on the prevalence of hospital infections and antimicrobial resistance in Kosova are outlined.

Total and attributable costs of surgical-wound infections at a Canadian tertiary-care center.

OBJECTIVE: To determine the total and attributable costs of surgical-wound infections in a Canadian teaching hospital. DESIGN: Retrospective incidence series study with chart review and examination of resource utilization attributable to wound infection. The charts of inpatients with wound infections were examined using the Appropriateness Evaluation Protocol (AEP), a validated chart review instrument designed to determine appropriateness of care, modified for wound infections. SETTING: A university referral center in Canada. PATIENTS: Medical records were abstracted from patients with wound infections who underwent an inpatient clean or clean-contaminated procedure during 1991. MEASUREMENTS: During the wound-infection treatment period, the hospital costs associated with providing care were tabulated for all inpatient days and for outpatient and emergency visits. Costs taken into account included nursing salary and benefits, nonphysician professional services, operating room time, laboratory, pharmacy, supplies, ancillary tests, and hotel costs. RESULTS: We identified 108 wound infections. Twenty-two patients required 28 surgical procedures related to a wound infection. Inpatient days totalled 1,116, costing $394,337. Fifty-five emergency and 42 clinic visits occurred, costing $27,193. By applying the AEP to the inpatient days, 833 days, or 10.2 days per case, were directly attributable to the wound infection. The hospital costs for inpatient care attributable to wound infections were $321,533 in total, or $3,937 per infection. Costs were distributed as follows: nursing, 51%; hotel, 14%; pharmacy, 10%; laboratory, 9%; emergency and outpatient clinic, 6%; professional services, 5%; operating room, 3%; and ancillary tests, 2%. CONCLUSIONS: Wound infections contribute markedly to extra days of hospitalization and related costs. The AEP method is applied easily to determine attributable days of care and costs of wound infections, which are necessary to calculate the cost-benefit of infection control programs.

A Canadian survey of prophylactic antibiotic use among hip-fracture patients.

OBJECTIVE: To study how surgical prophylactic antibiotics (SPAs) were utilized in the perioperative management of surgery for hip fractures. DESIGN: Retrospective chart review of randomly selected medical records. SETTING: Twenty-two hospitals (teaching, nonteaching, community, and large urban referral centers) from across Canada. PATIENTS: Patients admitted in 1990 with a diagnosis of hip fracture. METHODS: Complete medical records of 438 patients were examined; 352 cases who underwent surgical repair of a fractured hip with insertion of prosthetic material were included in analysis. Perioperative SPA use was assessed by abstracting the agent(s) chosen, dosages, time given with respect to the incision, and duration of postoperative use. Fourteen patient and process-of-care variables related to SPA were examined. RESULTS: 247 (70%) of 352 cases did not receive a dose of SPA 2 hours preoperatively. Ten percent of preoperative SPA was administered either too early or during the procedure. In 91 (39%) of 231 cases receiving SPA, the first dose was not administered until the end of the procedure. Preoperative SPA consisted of a parenteral first-generation cephalosporin for 94% of cases. SPAs were continued more than 24 hours postoperatively in 78% of cases. Lack of a written order for SPA, being a nonteaching hospital, and shorter duration of surgical procedure were predictive of failure to receive SPA in an effective manner. CONCLUSIONS: Most hip-fracture-surgery patients did not receive effective antibiotic prophylaxis as required to prevent serious wound infections. This important variable can be included for surveillance, so that corrective measures can be taken to assure effective prophylactic antibiotic administration.

Surgical wound infections occurring in day surgery patients.

The occurrence of surgical wound infection in outpatient day surgery has not been extensively studied despite the increasing popularity of this mode of treatment. The present study was conducted to determine the frequency of surgical wound infections in a day surgery population. We randomly selected during a 6-month period 635 (25%) of 2540 patients undergoing a day surgery procedure in which a skin incision was made. The patients were telephoned 1 month after their procedure by an infection control practitioner. Infection was diagnosed if the patient reported that (1) their physician had made a diagnosis of a wound infection or (2) pus was or had been issuing from the wound. Of the 515 patients contacted, 72% had undergone a clean and 28% a clean-contaminated procedure. Patient risk factors for infection were almost completely absent in our day surgery patients. Twenty-six wound infections were diagnosed, 19 of which were identified by physicians' diagnosis and 7 by patient description, for a rate of 5.05%. Two patients required hospitalization for their infections, and 14 were treated with antibiotics. The clean wound infection rates were 4.62%, less than half the infection rate seen in our patients undergoing inpatient surgery at 1 month follow-up by the same surveillance technique. We conclude that day surgery infection rates are much lower than inpatient surgery infection rates at our facility, probably because of a relative absence of risk factors in the day surgery patients.

Analysis of nanomolar S-nitrosothiol concentrations in physiological media.

S-Nitrosothiols occur endogenously and are thought to function as storage forms and/or stable carriers of nitric oxide. Moreover, the S-nitrosothiols have been postulated to function as neurotransmitters and mediate the vasodilator action of glyceryl trinitrate. Because of the increasing pharmacological and physiological interest in S-nitrosothiols, a sensitive method for analysis of these substances is required. We describe a sensitive method based on adsorptive stripping voltammetry for measurement of two S-nitrosothiols, namely, S-nitroso-N-acetyl-D-penicillamine (SNAP) and S-nitrosoglutathione (SNOG), in Krebs' solution. The method is based on the irreversible electrochemical reduction of SNAP and SNOG at the hanging mercury drop electrode (HMDE). The analytes were adsorbed at the HMDE for 60 s at -0.100 V, then exposed to a cathodic linear potential scan of 100 mV s(-10) which resulted in the reduction of SNAP at -0.470 V and SNOG at -0.530 V. Under these conditions, 4 nM SNAP and 11 nM SNOG were readily quantified. Using the above method, we were able to confirm the rapid degradation of SNAP under UV irradiation. Reproducibility of the method as applied to the analysis of these S-nitrosothiols in Krebs' solution was demonstrated by the within-day and day-to-day coefficients of variation of 1.5% and 2.0%, respectively.

Analysis of pyocyanin from Pseudomonas aeruginosa by adsorptive stripping voltammetry.

We have developed an adsorptive stripping voltammetric (AdSV) method that can be used for analysis of pyocyanin at the nanomolar level. The method employs a hanging mercury drop electrode (HMDE) as a working electrode in a three-electrode system in a 15-mL quartz cell. Preconcentration of pyocyanin in a 10-mM ammonia buffer (pH 8.0) on the HMDE is carried out by adsorption under conditions of controlled mass transfer for 60 sec, followed by scanning the electrode potential that results in reduction (cathodic scan) or oxidation (anodic scan) of the accumulated pyocyanin. For analysis of pyocyanin in samples containing a significant amount of surface-active species from a bacterial culture broth, a positive-going (anodic) potential scan must be applied after an adsorption at -0.50 V. Anodic scan of the adsorbed pyocyanin provided a well-defined oxidation peak at a potential of -0.17 V. The "anodic scan" variant of the method was tested for utility in Mueller-Hinton broth diluted from 20 to 200 times in a 10-mM ammonia buffer. Reproducibility of the method as applied to the analysis of pyocyanin produced by Pseudomonas aeruginosa (18.83 +/- 0.32 microM) in culture was demonstrated by the within-day and day-to-day coefficients of variation of 1.7% and 3.4%, respectively.

Growth of coagulase negative staphylococci in peritoneal dialysate fluid. Effect of calcium concentration.

The growth of coagulase negative staphylococci (CNS) in spent and unspent peritoneal dialysate of physiologic (1.25 mmol/l) and standard (1.62 mmol/l) [Ca2++] was studied. Dialysate was prepared by filter sterilization and pH and iCa were measured. Three isolates of CNS grown in unsupplemented Mueller Hinton broth for 18 to 20 hr at 37 degrees C were adjusted to a density of 1.5 x 10(8) CFU/ml. The dialysate (5.4 ml) was inoculated with 0.6 ml of a 10(-4) dilution and then incubated aerobically at 35 degrees C. Plates were made at 0, 6, and 24 hr, and CFU/100 microliters were counted after 24 hr. In a larger study, 10 isolates of CNS were examined. Coagulase negative staphylococcal growth was inhibited by unspent dialysate (P = 0.000; unpaired t-test). Growth was greater in spent physiologic [Ca++] dialysate (iCa -0.76 mmol/l) at 6 hr (P = 0.007) and at 24 hr (P = 0.000) than in standard dialysate (iCa = 0.86 mmol/l) for two of three strains in the pilot study. Similar findings were noted for 4 of 8 strains in the larger study. There was an overall trend for greater growth in physiologic [Ca++] dialysate (P = 0.09). Some strains of CNS grow better in lower [Ca++] dialysate, perhaps representing clinically relevant trophism.

Mycetoma of the foot caused by Cylindrocarpon destructans.

A 39-year-old male, originally from Antigua, West Indies, presented with a 12-year history of swelling of the left foot. A pathogen could not be recovered in cultures of three surgical biopsy specimens. During follow-up, pus and grains were expressed from a draining sinus tract and Cylindrocarpon destructans grew in pure culture. Retrospective examination of histologic sections of tissue removed during the third biopsy demonstrated a grain characteristic of eumycotic mycetoma. Although the fungus was susceptible to amphotericin B and ketoconazole in vitro, the patient refused treatment, and the clinical course over almost 19 years has been one of slow but progressive bone destruction. The fungus was identified by its microconidial morphology, the presence of chlamydospores, and an intense brown diffusible pigment. It was compared with another poorly known agent of white grain mycetoma, Phialophora cyanescens, characterized by phialidic conidia, chlamydospores in aggregations, and an intense diffusing pigment. The new combination Cylindrocarpon cyanescens (de Vries et al.) Sigler comb. nov. is proposed.

Fatal falciparum malaria in Canadian travellers.

The authors report 2 cases of severe falciparum malaria in Canadians that had fatal outcomes. In the first case a man presented to a local hospital shortly after returning from Africa, but a diagnosis of malaria was not considered. He was transferred to a secondary and then to a tertiary care facility, where he subsequently died. Intravenous quinidine therapy, the treatment of choice, was unavailable at all 3 hospitals. In the second case, a woman taking chloroquine prophylaxis while visiting Nigeria developed cerebral malaria and died. These cases illustrate critical management issues: appropriate advice on malaria prevention before departure; consideration of malaria in all febrile people returning from an endemic area; ready access to parenteral therapy for severe malaria in Canadian hospitals; and an increase in awareness of travel medicine among family physicians.

Unsuspected dilution of glutaraldehyde in an automatic washer for flexible fibreoptic endoscopes.

It has been suggested that serial usage of automatic endoscope washer-disinfectors may result in dilution of glutaraldehyde (used as the disinfectant in these machines). However, there are no published data available to confirm this possibility. The authors therefore undertook a study to determine the extent of glutaraldehyde dilution occurring during repeated cycles of an endoscope washer-disinfector over a 14 day period. It was found that the concentration of glutaraldehyde in the endoscope washer-disinfector consistently fell with increasing numbers of wash cycles, and that after 60 cycles the concentration of glutaraldehyde fell below the recommended minimum effective concentration of 1%. Users of automatic endoscope washer-disinfectors should be aware of the potential for dilution of glutaraldehyde in these machines. It is the number of washer-disinfector cycles, and not only the elapsed time since activation of the glutaraldehyde, which determines the frequency with which the glutaraldehyde solution must be replenished. Test kits are now available to measure glutaraldehyde concentration chemically, and may be useful in a quality control program of serial testing of glutaraldehyde concentrations in the endoscope washer-disinfector.

Granulomatous hepatitis and fever of unknown origin. An 11-year experience of 23 cases with three years' follow-up.

Granulomatous hepatitis is a common cause of fever of unknown origin in up to 13% of patients with prolonged fever. Attempts to define an exact etiology of the granulomatous hepatitis frequently does not yield a precise diagnosis, so that the physician must consider empiric treatment. In this paper we retrospectively review 23 patients in whom granulomatous hepatitis was found as part of the initial assessment of fever of unknown origin, and we report on their outcomes after an overall prospective follow-up of 37 months. In 26% a precise diagnosis was established at the time of assessment: Q-fever in three, mycobacterial disease in two, and histoplasmosis in one. In the remaining 74% no etiology was established after 44 months follow-up. Forty-one percent of the idiopathic group resolved spontaneously without therapy, and 18% received short-term prednisone or indomethacin with a favourable outcome. The remaining 41% required long-term prednisone therapy for a mean of 33.1 months, but all have remained afebrile and otherwise healthy after 59.6 months follow-up. We conclude that patients with fever of unknown origin who are diagnosed as having idiopathic granulomatous hepatitis have an excellent prognosis, even the minority who require long-term corticosteroids.

Antimicrobial activity of lidocaine against bacteria associated with nosocomial wound infection.

The authors characterized the in vitro antibacterial properties of clinical doses of lidocaine on isolates of a variety of bacterial pathogens commonly encountered in the setting of nosocomial wound infection (Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus) as well as a number of resistant strains of methicillin-resistant S. aureus and vancomycin-resistant enterococci. Time-kill studies were carried out on bacteria exposed to various clinical concentrations of lidocaine (0%, 1%, 2%, and 4%) with and without epinephrine (1:100,000). Minimum inhibitory concentrations and minimum bactericidal concentrations were determined for some strains using a broth macrodilution method recommended by the National Committee of Clinical Laboratory Standards. Lidocaine demonstrated a dose-dependent inhibition of growth for all strains of bacteria tested. The greatest sensitivity to lidocaine was shown by gram-negative organisms; the least sensitive was S. aureus. The addition of epinephrine to the local anesthetic had no effect on the susceptibility of the bacteria to lidocaine. These observations suggest that the surgical benefit of local anesthesia may extend beyond its analgesic properties and may have a role in the prophylaxis and, in the case of methicillin- and vancomycin-resistant bacteria, the treatment of surgical wound infection, mandating a wider application of this modality.

Electrospray mass-spectrometric, spectrophotometric and electrochemical methods do not provide evidence for the binding of nitric oxide by pyocyanine at pH 7.

In several recent publications on pyocyanine, its mechanism of action has been attributed to an ability to react with nitric oxide (NO), resulting in the formation of an adduct. We examined the chemical interaction of pyocyanine and NO using electrospray (ES) MS, spectrophotometry and voltammetry at neutral pH and with 10-100 microM pyocyanine. No binding of NO to pyocyanine was observed. Alternative mechanisms for the inhibition of NO-induced vasorelaxation by pyocyanine should be sought.

The role of polar pili in the adherence of Pseudomonas aeruginosa to injured canine tracheal cells: a semiquantitative morphologic study.

Pseudomonas aeruginosa adheres to respiratory epithelial cells in a highly specific fashion. In order to study the role of P. aeruginosa polar pili in the adherence process we conducted a quantitative morphological electron microscopic examination of P. aeruginosa adherence to SO2 injured canine tracheal cells in vitro. A pilin lacking background strain of P. aeruginosa PAK (BLP2) was constructed using a gene replacement and it in turn was engineered to express either the pilin gene of P. aeruginosa PAO, PAK, or no pilin gene. After 30 minutes incubation of these bacterial strains with injured canine tracheal rings the P. aeruginosa strains expressing pili adhered quantitatively more to the injured tracheal cells than did the pili lacking strains. PAO bearing strains adhered in greater numbers than PAK bearing strains. Healthy tracheal cells did not have any bacteria bound to their surfaces. The bacteria bound to the cilia and lateral edge of the exfoliating tracheal cells. Invasion of tracheal cells by piliated P. aeruginosa bacteria and penetration into the submucosa was also demonstrated. These data confirm the role of pili as important adhesins to injured tracheal cells. The difference in the adherence characteristics of pilin types PAK versus PAO may relate to the differences in the primary structure of these two pilin molecules.