Franz-Josef Schmitz

Molecular Detection of Antimicrobial Resistance

SUMMARY The determination of antimicrobial susceptibility of a clinical isolate, especially with increasing resistance, is often crucial for the optimal antimicrobial therapy of infected patients. Nucleic acid-based assays for the detection of resistance may offer advantages over phenotypic assays. Examples are the detection of the methicillin resistance-encoding mecA gene in staphylococci, rifampin resistance in Mycobacterium tuberculosis, and the spread of resistance determinants across the globe. However, molecular assays for the detection of resistance have a number of limitations. New resistance mechanisms may be missed, and in some cases the number of different genes makes generating an assay too costly to compete with phenotypic assays. In addition, proper quality control for molecular assays poses a problem for many laboratories, and this results in questionable results at best. The development of new molecular techniques, e.g., PCR using molecular beacons and DNA chips, expands the possibilities for monitoring resistance. Although molecular techniques for the detection of antimicrobial resistance clearly are winning a place in routine diagnostics, phenotypic assays are still the method of choice for most resistance determinations. In this review, we describe the applications of molecular techniques for the detection of antimicrobial resistance and the current state of the art.

Characterization of grlA,grlB, gyrA, and gyrB Mutations in 116 Unrelated Isolates of Staphylococcus aureus and Effects of Mutations on Ciprofloxacin MIC

ABSTRACT One hundred sixteen unrelated clinical isolates ofStaphylococcus aureus (70 ciprofloxacin resistant and 46 ciprofloxacin susceptible) from eight countries were studied for the presence of mutations in the grlA, grlB,gyrA, and gyrB gene loci. Two mutations withingrlA (located at codons 80 and 84) and two mutations withingyrA (located at codons 84 and 88) were clearly associated with ciprofloxacin resistance, although other mutations detected within the four genes studied may also contribute to decreased susceptibility.

Molecular surveillance of macrolide, tetracycline and quinolone resistance mechanisms in 1191 clinical European Streptococcus pneumoniae isolates

Streptococcus pneumoniae isolates (n=1191) were collected during a 1997-1999 European surveillance study. In addition to susceptibility data, a molecular epidemiological survey of their mechanisms of resistance to macrolides, tetracyclines, and quinolones was provided. Of the isolates tested, 72.6% were penicillin-susceptible, 19.9% penicillin-intermediate and 7.5% penicillin-resistant. There was an obvious relationship between resistance to penicillin and resistance to erythromycin (19% of all isolates), clindamycin (14%) and tetracycline (23%). Only one isolate was resistant to levofloxacin. Seventy-three percent of the European S. pneumoniae isolates resistant to erythromycin (n=229) carried the erm(B) gene, while the remaining 27% possessed the mef(A) gene. No mutations were detected in 23S rRNA or in ribosomal proteins L4 and L22. All tetracycline-resistant isolates (n=277) carried the tet(M) gene; none carried the tet(O) gene. Classical mutations in gyrA (Ser 81-Phe or Tyr) and parC (Ser 79-Phe and Asp 83-Asn) and efflux contributed to the decreased quinolone susceptibility. This study of recent European S. pneumoniae isolates can be used to recognize any changes in susceptibility patterns and resistance mechanisms that may occur in the future.

Comparative activities of six different fluoroquinolones against 9,682 clinical bacterial isolates from 20 European university hospitals participating in the European SENTRY surveillance programme

The in-vitro activities of gatifloxacin, trovafloxacin, levofloxacin, sparfloxacin, ofloxacin, and ciprofloxacin were tested against 9,682 clinical bacterial isolates from 20 European university hospitals participating in the European SENTRY surveillance programme. Gatifloxacin and trovafloxacin exhibited the highest activities against gram-positive cocci, while levofloxacin, ofloxacin, ciprofloxacin, and gatifloxacin were the most active against Enterobacteriaceae. Ciprofloxacin and levofloxacin showed the highest antimicrobial activities against Pseudomonas spp., while gatifloxacin and trovafloxacin were the most active against Acinetobacter spp. and Stenotrophomonas maltophilia. All Haemophilus spp. and Moraxella catarrhalis isolates were fully susceptible to all quinolones tested. Overall, the new quinolones, showed improved activity against gram-positive cocci and gram-negative non-fermenters while retaining their broad-spectrum activity against gram-negative bacilli.