Glycopeptide-intermediate Staphylococcus aureus (GISA) and, in particular, heterogeneous GISA (hGISA) are difficult to detect by standard MIC methods, and thus, an accurate detection method for clinical practice and surveillances is needed. Two prototype Etest strips designed for hGISA/GISA resistance detection (GRD) were evaluated using a worldwide collection of hGISA/GISA strains covering the five major clonal lineages. A total of 150 strains comprising 15 GISA and 60 hGISA strains (defined by population analysis profiles-area under the curve [PAP-AUC]), 70 glycopeptide-susceptible S. aureus (GSSA) strains, and 5 S. aureus ATCC reference strains were tested. For standardized Etest vancomycin (VA) MIC testing, the modified Etest macromethod with VA and teicoplanin (TP) strips tested with a heavier inoculum using brain heart infusion agar (BHI) and two glycopeptide screening agar plates (6 g/ml VA/BHI and 5 g/ml Mueller-Hinton agar [MHA]) were tested in parallel with the two new Etest GRD strips: a VA 32 (0.5-g/ml)-TP 32 (0.5-g/ml) double-sided gradient (E-VA/TP) with one prototype overlaid with a nutrient (E-VA/TP؉S) to enhance the growth of hGISA. The Etest GRD strips were tested with a standard 0.5-McFarland standard inoculum using MHA and MHA plus 5% blood (MHB) and were read at 18 to 24 and 48 h. The interpretive MIC cutoffs used for the new Etest GRD strips at 24 and 48 h were as follows: for GISA, TP or VA, >8, and a standard VA MIC of >6; for hGISA, TP or VA, >8, and a standard VA MIC of <4. The results on MHB at 48 h showed that E-VA/TP؉S had high specificity (94%) and sensitivity (95%) in comparison to PAP-AUC and was able to detect all GISA (n ‫؍‬ 15) and 98% of hGISA (n ‫؍‬ 60) strains. In contrast, the glycopeptide screening plates performed poorly for hGISA. The new Etest GRD strip (E-VA/TP؉S), utilizing standard media and inocula, is a simple and acceptable tool for detection of hGISA/GISA for clinical and epidemiologic purposes.

Clinical microbiology has always been a slowly evolving and conservative science. The sub-field of bacteriology has been and still is dominated for over a century by culture-based technologies. The integration of serological and molecular methodologies during the seventies and eighties of the previous century took place relatively slowly and in a cumbersome fashion. When nucleic acid amplification technologies became available in the early nineties, the predicted "revolution" was again slow but in the end a real paradigm shift did take place. Several of the culture-based technologies were successfully replaced by tests aimed at nucleic acid detection. More recently a second revolution occurred. Mass spectrometry was introduced and broadly accepted as a new diagnostic gold standard for microbial species identification. Apparently, the diagnostic landscape is changing, albeit slowly, and the combination of newly identified infectious etiologies and the availability of innovative technologies has now opened new avenues for modernizing clinical microbiology. However, the improvement of microbial antibiotic susceptibility testing is still lagging behind. In this review we aim to sketch the most recent developments in laboratory-based clinical bacteriology and to provide an overview of emerging novel diagnostic approaches.

A multicenter study was conducted to validate Etest tigecycline compared to the Clinical Laboratory Standards Institute reference broth microdilution and agar dilution methodologies. A large collection of gram-negative (n ‫؍‬ 266) and gram-positive (n ‫؍‬ 162) aerobic bacteria, a collection of anaerobes (n ‫؍‬ 385), and selected collections of nonpneumococcal streptococci (n ‫؍‬ 369), Streptococcus pneumoniae (n ‫؍‬ 372), and Haemophilus influenzae (n ‫؍‬ 372) were tested. Strains with reduced susceptibility to tigecycline were used with all test methods. The Etest showed excellent inter-and intralaboratory reproducibility for all organism groups tested regardless of the test methodology. The essential agreement values with the reference method (؎1 dilution) were >99% for the collection of gram-negative and gram-positive aerobes; >98% for the S. pneumoniae, H. influenzae, and anaerobe collections; and 100% for the group of nonpneumococcal streptococci. These results validate the performance accuracy and utility of Etest tigecycline and verify the reproducibility of this convenient predefined gradient methodology for tigecycline susceptibility determination.

Thirty-five Bacteroides fragilis clinical isolates with varying susceptibility to meropenem were analysed with a prototype of a double-ended Etest strip containing meropenem +/- EDTA, designed for the detection of the CfiA metallo-beta-lactamase. Phenotypic results obtained with this new Etest strip were related to the genotype and compared to the results of the Etest containing imipenem +/- EDTA. Whereas the Etest with imipenem +/- EDTA only allowed detection of isolates with high-level resistance (both MICs of imipenem and meropenem >32 mg/L), reflecting the possible underestimation of CfiA prevalence in B. fragilis, the Etest with meropenem +/- EDTA proved to be more accurate, particularly for isolates with low-level carbapenem resistance, suggesting its potential for broader detection of CfiA production.

The new test was shown to be diagnostically useful, easy to implement and less labour intensive than the classical method.