Matrix assisted laser desorption ionization time of flight mass spectrometry for microbial identification in clinical microbiology / 中华检验医学杂志

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) recently moved from physics laboratories into diagnostic clinical microbiology.In this capacity it is currently replacing phenotypic bacterial identification tools that were designed and developed by Pasteur and Koch.Many European microbiology laboratories now use MALDI-TOF-MS with a high degree of efficiency.In the USA this will soon become common as well since the Food and Drug Administration (FDA) has now cleared the first of the commercially available systems for in vitro diagnostic use.This will also further spark the development of mass spectrometry applications in the field of antimicrobial susceptibility testing and epidemiological typing.This review summarizes the current state of affairs with respect to MALDI-TOF-MS in clinical microbiology.Emphasis will be on clinical relevance and studies in that field and the diagnostic data obtained through comparative analyses of different MALDI-TOF-MS instrumentation and through multi-center validation studies will be reviewed.

Rapid inactivation of Mycobacterium and nocardia species before identification using matrix-assisted laser desorption ionization-time of flight mass spectrometry.

The identification of mycobacteria outside biocontainment facilities requires that the organisms first be rendered inactive. Exposure to 70% ethanol (EtOH) either before or after mechanical disruption was evaluated in order to establish a safe, effective, and rapid inactivation protocol that is compatible with identification of Mycobacterium and Nocardia species using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). A combination of 5 min of bead beating in 70% EtOH followed by a 10-min room temperature incubation period was found to be rapidly bactericidal and provided high-quality spectra compared to spectra obtained directly from growth on solid media. The age of the culture, the stability of the refrigerated or frozen lysates, and freeze-thaw cycles did not adversely impact the quality of the spectra or the identification obtained.

Prospective evaluation of the Meridian Illumigene™ loop-mediated amplification assay and the Gen Probe ProGastro™ Cd polymerase chain reaction assay for the direct detection of toxigenic Clostridium difficile from fecal samples.

Clostridium difficile is the most common and important cause of toxigenic colitis in the health care setting. Laboratory diagnostics have included bacterial culture with further identification of toxigenic stains, or more commonly, direct detection of preformed toxin in stool samples using biological or immunochemistry assays. Recently, molecular amplification assays for the direct detection of toxin-encoding genes have become available commercially. We prospectively evaluated 2 FDA-cleared molecular amplification tests, the Illumigene C. difficile and the ProGastro Cd PCR assay, for the direct detection of toxigenic C. difficile from fecal samples. Of 446 samples tested, 418 produced matching amplification results, 88 positive and 330 negative, and 13 resolved with repeat testing. Toxigenic culture and direct cytotoxin testing were used to resolve the remaining 15 discordant samples. Overall, each assay performed well and correctly identified 97% of positive samples.

Reevaluation of the Premier Clostridium difficile toxin A and B immunoassay with comparison to glutamate dehydrogenase common antigen testing evaluating Bartels cytotoxin and Prodesse ProGastro Cd polymerase chain reaction as confirmatory procedures.

Enzyme immunoassays are currently the most common tests used in the clinical laboratory for the detection of Clostridium difficile toxins; however, significant problems with their performance have recently been described. We prospectively reevaluated the Meridian Premier C. difficile toxin A/B assay with direct comparison to a 2-step algorithm that screened for C. difficile common antigen and compared cytotoxin and real-time polymerase chain reaction (PCR) as confirmatory procedures. The Premier assay lacked sufficient sensitivity, missing 25% of true-positive samples. PCR was the most sensitive method and the only procedure that allowed same day testing and reporting.

Shiga toxin-producing Escherichia coli as a possible etiological agent of chronic diarrhea.

We identified Shiga toxin-producing Escherichia coli (STEC) as the likely etiologic pathogen for chronic diarrhea in 2 patients, 1 of whom was immunocompromised with acquired immunodeficiency syndrome, and 1 of whom was immunocompetent. Both were treated with antibiotics, and neither developed systemic complications of the infection. These cases suggest that STEC infection should be considered in the differential diagnosis of chronic diarrhea.

Prospective comparison of the Gen-probe PACE 2 assay and the Abbott ligase chain reaction for the direct detection of Chlamydia trachomatis in a low prevalence population.

In a prospective study, the Gen-Probe PACE 2 (GP) assay was compared with Abbott Laboratories' ligase chain reaction (LCR) assay for the detection of Chlamydia trachomatis. A total of 493 female patients consented to collection of two cervical samples; a first-void urine (FVU) sample was collected also from 446 of the participants. Cervical samples were tested by both GP and LCR; 16 samples (3.1%) tested positive by both methods and no discrepant results were observed. All but one of the FVU samples collected from patients with a positive cervical sample was positive for C. trachomatis by LCR. The stability of FVU samples over time in the LCR test was also evaluated and proved to be significantly longer than the 4 days stated by the manufacturer. While LCR proved to be highly sensitive in detecting chlamydial infection in FVU samples, no difference was noted between LCR and GP in the detection of cervical C. trachomatis infection in this study population.