A comparative study was designed to evaluate the identification (ID) and antimicrobial susceptibility testing (AST) performances of the BD Phoenix Automated Microbiology System (Becton Dickinson Diagnostic Systems [BD], Pont de Claix, France). A total of 305 single clinical isolates were collected, and comparisons were made with routine manual methods in use in our microbiology laboratories. The percentages of correct IDs were 93.3, 89.4, 91.8, and 85.7% for enterobacteria, nonfermenting gram-negative bacilli, staphylococci, and streptococci-enterococci, respectively. The median ID time was 3 h, and the median time for AST was 10 h 30 min. AST results showed variable percentages of errors for the different antibiotics. None of the enterobacteria and 0.3% of Pseudomonas aeruginosa isolates showed a very major error (VME). Only one strain of Staphylococcus aureus showed a VME with oxacillin. We demonstrate here the efficiency of the Phoenix system, which can be used for the majority of strains encountered in a university-based laboratory, for ID and AST.Automation in microbiology is not an easy task because microbiology is still a discipline that requires input from laboratory staff for the interpretation of results. This is specifically the case for bacterial identification (ID) and detection of antibiotic resistance, since microbiologists are often able to identify a bacterium and its resistance phenotype on the basis of only a few parameters. However, biochemical ID and susceptibility testing require 24 h to be readable. Several factors favor the use of automatic systems in the microbiology laboratory. Reproducibility, the ability to track results, the availability of results within one working day, and reduced amounts of contaminated waste represent the main reasons. An added advantage includes an automatic connection to the laboratory informatics software, allowing better management of technician staff, an easier validation for the microbiologist with the help of an expert system, and the opportunity for the clinician to obtain partial or complete results faster, thus improving patient management. Over the last 20 years, a variety of automated systems for ID and antimicrobial susceptibility testing (AST) have been developed (12,13,16,17,27,33,35,(36)(37)(38)(39)(40)(41)43). These automated technologies allowed for the very first time complete management of ID and AST, requiring technical staff to perform only one dilution from an agar plate culture. For example, the Vitek 2 (bioMérieux, Marcy l'Étoile, France) and the BD Phoenix (Becton Dickinson Diagnostic Systems [BD], Pont de Claix, France) are new systems that automatically perform ID and AST on a manually prepared inoculum (1, 3, 6, 9, 10, 14, 15, 18, 19, 22-25, 31, 42). Both designs possess an advanced expert system and have a potential impact on the workflow of a clinical laboratory for a typical hospital. Moreover, rapid ID and AST can have a significant impact on the management of infections, especially those caused by antibiotic-resistant bacteria ...
Granzyme B serine protease is found in the granules of activated cytotoxic T cells and in natural and lymphokine-activated killer cells. This protease plays a critical role in the rapid induction of target cell DNA fragmentation. The DNA regulatory elements that are responsible for the specificity of granzyme B gene transcription in activated T-cells reside between nt -148 and +60 (relative to the transcription start point at +1) of the human granzyme B gene promoter. This region contains binding sites for the transcription factors Ikaros, CBF, Ets, and AP-1. Mutational analysis of the human granzyme B promoter reveals that the Ikaros binding site (-143 to -114) and the AP-1/CBF binding site (-103 to -77) are essential for the activation of transcription in phytohemagglutinin-activated peripheral blood lymphocytes, whereas mutation of the Ets binding site does not affect promoter activity in these cells.
We have investigated perforin and granzyme B expression in graft-infiltrating lymphocytes of patients who underwent heart transplantation. Those proteins are commonly present in the cytoplasmic granules of cytotoxic T lymphocytes and are released upon effector-target cell interaction. From 28 patients 103 endomyocardial biopsies were obtained and examined by histology and immunocytochemical analysis using relevant monoclonal antibodies. We found that "high" biopsy histological grades were associated with perforin and granzyme B expression in graft-infiltrating lymphocytes of patients with acute severe rejection crisis. In contrast, these markers were not detected in patients without rejection or during graft stabilization. Interestingly, in patients with mild rejection and "low" histological grades, two groups could be distinguished with a differential expression of the two intracytoplasmic proteins. The presence of perforin and granzyme B-expressing cells was found to be predictive of rapid progression to severe rejection, so that this situation required additional treatment; in contrast, their absence seemed to correlate with a good graft outcome without additional treatment. Moreover, perforin and granzyme B expression seemed to be down-regulated by immunosuppressive drugs, which coincided with graft stabilization. In conclusion, our data suggest that detection of granzyme B and perforin in graft-infiltrating lymphocytes might be helpful for routinely monitoring heart transplant patients.
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