Oxacillin Information and Courses from MediaLab, Inc.
These are the MediaLab courses that cover Oxacillin and links to relevant pages within the course.
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|The zone of inhibition around the disk shown in the image has been measured at 23 mm. Based on this result, you should:||View Page|
|The oxacillin screen test alone is not sufficient for determining the susceptibility to penicillin for S. pneumoniae isolates recovered from cerebrospinal fluid (CSF).||View Page|
Most S. pneumoniae strains gain penicillin resistance by altering the penicillin-binding proteins in their cell wall. Penicillin molecules that cannot find a penicillin binding site cannot interfere with cell wall synthesis. Several different types of penicillin binding proteins may be involved, explaining the various levels of intermediate resistance that may be encountered with different strains of S. pneumoniae. Because different penicillin binding proteins may be involved, the level of penicillin resistance cannot be predicted by the oxacillin screening test. Infections caused by isolates of S. pneumoniae showing penicillin resistance in the intermediate range may be successfully treated by administering high doses of antibiotic. For this reason, the level of resistance with an accurate minimum inhibitory concentration (MIC) test must be determined for all clinically significant isolates of S. pneumoniae.
|Methicillin-Resistant Staphylococcus aureus (MRSA) Screen|
Perhaps the most efficient means for detecting methicillin-resistant staphylococci in clinical laboratories is the use of the agar dilution screening test. Illustrated in the image is a Mueller-Hinton agar plate containing 6 ug/mL of oxicillin, previously inoculated with a strain of Staphylococcus aureus. Oxacillin is used as a marker for methicillin resistance because it is more stable in the agar medium. Growth on this screening medium is presumptive for methicillin resistance. Thus, in the presence of growth, as shown here, a follow-up minimum inhibitory concentration (MIC) test must be performed to determine the exact level of resistance.
|What is successful molecular identification of methicillin-resistant Staphylococcus aureus (MRSA) based upon? (Choose the BEST answer)||View Page|
|Beta-lactam antibiotics and S. aureus|
Antibiotics inhibit bacterial growth by interfering with one or more cellular processes. Beta lactams are a large group of cell wall active antibiotics used to treat a wide variety of infections. S. aureus cell wall synthesis is dependent on the proper functioning of a number of enzymes. The beta-lactam antibiotics exert their effect by binding with one specific type of enzyme, transpeptidase, thus interfering with its ability to catalyze the final stage of peptidoglycan synthesis, resulting in defective cell wall formation. The beta-lactams comprise four main groups of antibiotics; all have the beta-lactam ring as their basic chemical structure: Penicillins (penicillin, oxacillin/methicillin, ampicillin and piperacillin) Cephalosporins Carbapenems Monobactams The spectrum of antimicrobial activity is dependent upon the particular structural modification of the beta-lactam ring. The transpeptidases are commonly referred to as penicillin-binding proteins (PBPs). Different bacterial species have distinct PBPs, resulting in very specific drug interactions.
|Beta-lactams and Methicillin Resistant Staphylococcus aureus|
Methicillin Resistant Staphylococcus aureus (MRSA) is resistant to the beta-lactam antibiotics. The term methicillin-resistant is historically used to describe resistance to any of this class of antimicrobials even though methicillin is no longer the drug of choice. The acronym MRSA persists and is used interchangeably with ORSA – oxacillin-resistant Staphylococcus aureus. Oxacillin/methicillin resistance implies resistance to all penicillins, cephalosporins, monobactams, carbepenems and beta-lactam/beta-lactamase inhibitor combinations. S. aureus intrinsically produces beta lactamase enzymes that breakdown beta lactam antibiotics (i.e., penicillin); these are designated PBP 1 - 4. The beta-lactam resistance of MRSA is determined by the production of a novel penicillin binding protein called PBP 2' (PBP 2a), that has a reduced binding affinity for beta-lactam antibiotics. This allows MRSA strains to continue cell wall synthesis due to the uninhibited activity of PBP2' even in the presence of otherwise inhibitory concentrations of beta-lactam antibiotics.PBP2' is encoded by a mecA gene located on the MRSA chromosome and is widely distributed among Staphylococcus aureus as well as coagulase-negative staphylococci. The mecA gene is carried by a novel mobile genetic element, designated staphylococcal cassette chromosome mec – SCCmec that is integrated into the bacterial chromosome. The mecA gene is believed to have originated in some coagulase-negative staphylococcal strains and was then transferred into S. aureus, giving rise to MRSA. It is likely that SCCmec serves as the carrier of the mecA gene moving across staphylococcal spp. as these mecA genes have never been found without the presence of a SCCmec-like structure. Phylogenetic analyses of international collections of MRSA and methicillin-susceptible S. aureus isolates have revealed that methicillin resistance has arisen in five distinct lineages designated SCCmec I – V, which differ in both size and genetic composition. In recent years, the gene has continued to evolve so that many MRSA strains are currently resistant to several different antibiotics.
|Assume you perform microbiology for an institution submitting surveillance cultures for MRSA. Which isolate should receive further workup to rule out methicillin (oxacillin) resistance?||View Page|
|Your laboratory's primary susceptibility testing method is disk diffusion. The cefoxitin disc has a zone size of 19 mm and the vancomycin disc has a zone size of 7 mm. Appropriate courses of action include:||View Page|
When a clinical isolate is presumptively identified as S. aureus, susceptibility testing will be performed by either the Standardized Disk Diffusion (Kirby-Bauer) or Broth Dilution (MIC) methods, using the following testing conditions as recommended by the Clinical and Laboratory Standards Institute (CLSI): Medium: MHA for disk diffusion; CAMHB + 2% NaCL for oxacillin, methicillin, and nafcillin; CAMHB supplemented up to 50 ug/ml calcium for daptomycin Inoculum: Direct colony suspension (0.5 McFarland Standard) Incubation: 35° C (Testing at temperatures above 35° C may not detect MRSA); ambient air; disk diffusion; 16to 18 hours; dilution methods; 16 to 20 hours. All methods: 24 hrs for oxacillin, methicillin, nafcillin, and vancomycin.
|Interpretation of Oxacillin and Cefoxitin Disk Diffusion Tests|
Oxacillin is the agent of choice for standardized MIC methods (broth & agar dilution). However, since 2006 the Clinical Laboratory Standards Institute (CLSI) has recommended the use of 30 µg cefoxitin disk rather than the oxacillin disk to detect mecA-mediated resistance in the disk diffusion test because the cefoxitin disk test is easier to read and is as sensitive and specific as MIC methods. Results are still reported as "oxacillin-resistant" or "oxacillin-sensitive." Cefoxitin is a better inducer of the mecA gene and gives clearer, easier to read endpoints in disk diffusion tests.The oxacillin disk is read for light growth within the zone of inhibition using transmitted light (plate held up to light), ANY discernible growth within zone of inhibition is indicative of resistance. The cefoxitin disk is read using reflected light.Interpretive Critieria for Cefoxitin Disk Diffusion Test Resisitant Intermediate Susceptible S.aureus/MRSA < 21 mm N/A > 22 mm
|Detection of Oxacillin Resistance|
Resistance to oxacillin is most accurately determined by testing for mecA or for the protein expressed by mecA, the penicillin-binding protein 2a (PBP 2a, which is also referred to as PBP 2'). Isolates of staphylococci that carry the mecA gene, or that produce PBP 2a should be reported as oxacillin-resistant according to CLSI guidelines. If MIC tests are performed in addition to disk diffusion, isolates for which oxacillin MICs are > 4µg/mL, are mecA-negative, or PBP 2a negative should be reported as oxacillin-resistant. Such isolates may have a rare resistance mechanism other than mecA, and may also test susceptible to cefoxitin by disc diffusion. In these scenarios, oxacillin resistance should be reported in accordance with the MIC value.
|Interpretation of Oxacillin Broth Dilution Tests.|
The CLSI document M100 is the definitive resource for all standard susceptibility interpretative criteria.Oxacillin is the standard antibiotic incorporated into MIC testing algorithms to predict resistance to beta lactam antibiotics. Breakpoints for Oxacillin as defined by the CLSI are defined as follows: Interpretative criteria for Oxacillin and Staph aureus: Antibiotic Susceptible Intermediate Resistant Oxacillin < 2µg/mL -- > 4 µg/mL
|Which of the following scenarios represents appropriate detection of MRSA by the Kirby Bauer method?||View Page|
|Your laboratory performs MIC tests, in addition to Kirby Bauer and on blood culture isolates, a PBP 2a assay. Which of the following isolates should be reported as oxacillin resistant?||View Page|
As previously discussed, multiplexing is assaying several different genes in the same reaction. Multiplexing has become possible due the high specificity of probe design. Development of multiplexing PCR reactions can be complicated because it has to be ensured that the probes do not interfere or react with each other. The process can also be time-consuming because each reaction may require different conditions. Figuring out the ideal mix for all the probes can take time. When developing a new multiplex test, it is important to weigh the time involved and costs against running each PCR reaction individually. There are many pre-established multiplex tests that can be purchased and used in laboratories today. One example is a multiplex PCR test that can distinguish between Staphylococcus aureus and coagulase-negative Staphylococcus and determine the presence of the gene that is resistant to oxacillin. Another example is a multiplex PCR test that can detect all four species of Plasmodium, the cause of malaria.