Penicillinase testing is required for Staphylococcus aureus isolates with a penicillin MIC of

In total, 494 isolates of coagulase-negative staphylococci (CoNS) were identified to the species level by biochemical tests and sodA sequencing. Erythromycin resistance phenotypes were determined and specific resistance genes were identified by PCR. The prevalence of erythromycin resistance varied widely among staphylococcal species, from 0% in Staphylococcus lugdunensis to almost 90% in Staphylococcus haemolyticus. Most (63%) erythromycin-resistant isolates carried constitutively expressed erm(C) as the sole resistance determinant, with the notable exception of Staphylococcus hominis subsp. hominis, which carried inducible erm(C). The erm(A) and erm(B) determinants were comparatively rare. The msr(A) gene was carried by 20-30% of all erythromycin-resistant isolates, with little variation among species, and was combined in 16.7% of isolates with mph(C), a resistance gene of unknown clinical relevance found previously in isolates of veterinary origin. No erythromycin resistance that could not be attributed to the genes investigated was detected. It was concluded that the presence of methylases cannot be assumed in CoNS isolates that appear erythromycin-resistant and clindamycin-susceptible; thus, methods that detect the export mechanism should be used with clinically significant isolates to indicate whether use of clindamycin may be effective. In Staphylococcus epidermidis and S. haemolyticus, 46% and 66%, respectively, of erythromycin-resistant, clindamycin-susceptible isolates were susceptible to clindamycin therapy.

The human pathogen Helicobacter pylori has infected more than half of the world's population. Nevertheless, the first step of infection, the acute colonization of the gastric mucus, is poorly understood. For successful colonization, H. pylori must retain active motility in the gastric lumen until it reaches the safety of the mucus layer. To identify the factors determining the acute colonization, we inserted bacteria into the stomach of anesthetized Mongolian gerbils. We adjusted the gastric juice to defined pH values of between 2.0 and 6.0 by using an autotitrator. Despite the fact that Helicobacter spp. are known to survive low pH values for a certain time in vitro, the length of time that H. pylori persisted under the assay conditions within the gastric juice in vivo was remarkably shorter. In the anesthetized animal we found H. pylori to be irreversibly immotile in less than 1 min at lumen pH values of 2 and 3. At pH 4 motility was lost after 2 min. However, the period of motility increased to more than 15 min at pH 6. Blocking pepsins in the gastric lumen in vivo by using pepstatin significantly increased the period of motility. It was possible to simulate the rapid in vivo immotilization in vitro by adding pepsins. We conclude that pepsin limits the persistence of H. pylori in the gastric chymus to only a few minutes by rapidly inhibiting active motility. It is therefore likely that this short period of resistance in the gastric lumen is one of the most critical phases of Helicobacter infection.

Helicobacter pylori possesses a gene (HP0326/JHP309) homologous to neuA of other bacteria, encoding a cytidyl monophosphate-N-acetylneuraminic acid synthetase-homologous enzyme in its N-terminal portion. We analysed the function of this gene, which is controlled by a flagellar class 2 sigma(54) promoter, in flagellar biosynthesis. HP0326/JHP309 actually represents a bicistronic operon consisting of a neuA and a flmD-like putative glycosyl transferase gene. An isogenic flmD mutant synthesized basal bodies but no filaments, was non-motile, and expressed severely reduced amounts of a FlaA flagellin of reduced molecular mass. FlaA flagellin was found to be glycosylated in its exported form within the flagellar filament, but not inside the cytoplasm. Glycosylated FlaA was not detectable in the flmD mutant. Together with other genes in the H. pylori genome, a proposed function of the neuA/flmD gene products could be to provide a pathway for glycosylation of flagellin and other extracytoplasmic molecules during type III secretion.

Of 104 genotypically diverse methicillin-resistant Staphylococcus aureus (MRSA) isolates tested with the MicroScan WalkAway (Pos MIC 24 panel) and Vitek 2 (AST-P549 card) systems, 7 and 6 isolates, respectively, showed an oxacillin MIC of <2mg/liter. Most of these MRSA isolates were community acquired. However, if the cefoxitin screen of AST-P549 was also considered, MRSA detection failed for only one isolate.The prevalence of methicillin-resistant Staphylococcus aureus (MRSA) has increased over the last years. Reliable detection of MRSA is important since a false report of a patient's isolate as methicillin susceptible would result in inadequate therapy with probably fatal consequences (2). Whereas MRSA infections formerly occurred almost exclusively in hospitalized patients, community-acquired MRSA (cMRSA) isolates have been reported recently in patients without any previous contact with the health care system (7).Many laboratories rely on automatic susceptibility testing methods that use oxacillin MIC testing, oxacillin breakpoint detection in the presence of salt, or cefoxitin MIC testing as markers for the presence of methicillin resistance. Many studies have investigated the detection of MRSA by the Vitek 2 system (3,4,8,11,12,13,15,17); however, data for the performance of the MicroScan WalkAway system in MRSA detection are scarce (17).Most studies evaluating the performance of Vitek 2 used consecutive clinical strains (3,8,11,12,15), but this approach may be biased by the overrepresentation of locally predominant clones and may not predict performance in other geographical areas. We therefore used a collection of MRSA strains with distinct pulsed-field gel electrophoresis (PFGE) patterns to study MRSA detection using the MicroScan WalkAway and Vitek 2 systems.From 1998 to 2006, noncopy MRSA isolates (n ϭ 1,516), initially identified by oxacillin screening agar or Vitek, from four hospitals in the Bochum area were collected and typed by PFGE as described previously (5). Of these, 120 isolates with different PFGE patterns were chosen. The patterns were interpreted according to the criteria of Tenover et al. (18), and isolates grouped into PFGE types and subtypes.For susceptibility tests, isolates from frozen storage were subcultured twice on Columbia blood agar at 37°C in 5% CO 2 before being tested with the Vitek 2 system using the AST-P549 card and the MicroScan WalkAway system using the Pos MIC 24 panel.Whenever results for oxacillin in the Vitek 2 or MicroScan WalkAway system or for the cefoxitin screen in the Vitek 2 system were not indicative of MRSA, a mecA PCR was performed from colonies growing on purity control plates of both automatic systems and a S. aureus-specific PCR for SA442 (16) was used as an internal positive control. In addition, the Panton-Valentine leukocidin (PVL)-coding genes lukS-PVL and lukF-PVL were detected by PCR (9). SCCmec typing (10) and spa typing (6) were performed as described previously.Loss of mecA during storage of isolates could be demonstrated in 16 of 120 isolates ...