Department of Microbiology and Biomedical Sciences, University of Ancona Medical School, Via Ranieri, Monte dAgo, 60131 Ancona, Italy
Few subjects in clinical microbiology are debated as frequently and extensively as antimicrobial susceptibility testing. Nevertheless, few other subjects continue to be considered of such topical interest. Although this is owing to several factors, it mainly reflects on the one hand the continuous technical evolution in this field, and on the other the fact that all such innovations are substantially directed at better assessing antibiotic resistance, which is perceived as a major crisis area of modern medicine.
Little by little, yesterdays perhaps ingenuous hopes and the early dream of all-powerful antibiotics have been eroded and progressively replaced with deep distrust.1 The repeated warnings of microbial wars,2 new plagues,3 worldwide calamities,4 new apocalypses57 and even an impending post-antimicrobial era8 are a disquieting confirmation of this profound lack of confidence. It is a hard fact that antibiotic resistance is a growing phenomenon, whereas the rate of discovery of new antibiotics has been declining for years.9 In the late 1980s, many companies scaled down or stopped financing of antibiotic research or maintained only very focused programmes.1,10,11 This change in policy particularly affected the really new antibioticsi.e. those acting on totally new bacterial targetswhich, together with vaccine development, constitute the only hope of stemming the tide of drug resistance.1,9,11 Thus, whereas there always used to be a new antibiotic in our future, this is no longer the case.10 Spratt grimly argued that the bacteria will not give in, the drug companies will.1
Despite this, the use of antibiotics remains too high, sometimes as a form of consumerism fostered in turn by patients, physicians, chemists and drug companies. Alarmingly, however, mounting experimental evidence indicates that a reduction in the use of antibiotics and in the relevant selective pressure might not result in the disappearance of the resistant bacteria already present in hospitals and the community, and in human and environmental reservoirs.1214
Over the last few years, some new concepts on antimicrobial resistance have been emerging, particularly the relationship between resistance and pathogenicity. Courvalin15 has recently observed that for a bacterium it is better to be resistant than virulent, especially in a hospital setting. What is more, as a rule we only see the tip of the iceberg, as the reservoir for spread of antibiotic-resistant nosocomial pathogens usually consists of a relatively small number of patients with clinical evidence of infection and a much larger number of colonized patients.16 To these must be added animals and other reservoirs of nosocomial pathogens, as in the case of vancomycin-resistant enterococci, which cause infections that are currently regarded as a zoonosis.17 In the absence of active microbiological surveillance, these situations may go unnoticed and escape preventive measures. New and unexpected relationships between resistance and pathogenicity are also emerging among community pathogens. Facinelli et al.18 have recently shown in clinical isolates of Streptococcus pyogenes a close association between erythromycin resistance and ability to enter human respiratory cells. Hence, strains combining these two features may be able to escape the two major classes of antistreptococcal drugsß-lactams by virtue of intracellular location and macrolides by virtue of resistanceresulting in difficulty of eradication. This may have facilitated the wide and unparalleled diffusion of erythromycin-resistant S. pyogenes populations in Italy.1820
As regards antimicrobial susceptibility testing, the heart of the matter remains the tenuous, and sometimes ambiguous, link between in vitro and in vivo phenomena, i.e. between laboratory findings and their clinical relevance. The issue of the predictive value of in vitro susceptibility tests has perceptively been focused on by Phillips,21,22 who, in comparing the opinions of some healthily agnostic microbiologists, pioneered by Greenwood,23 with those of other researchers (the majority) with whom he sided, concluded that one can learn to accommodate most of the failings. It is well established that with susceptibility tests in vitro, the datum of resistance is more predictive than that of susceptibility.24 Indeed, if the laboratory test indicates that a clinical isolate is resistant in vitro to a particular antibiotic and that agent is used in therapy, the treatment is very likely to fail. But the converse is not necessarily true: if the laboratory test indicates that a clinical isolate is susceptible in vitro to a particular antibiotic, there is no guarantee that therapy with that agent will be successful, the clinical outcome depending on a wide range of other factors besides in vitro susceptibility (site of infection, pharmacological properties of the antibiotic, concomitance of other diseases, efficiency of specific and non-specific defence mechanisms, etc.). Thus, whereas in vitro resistance is a condition sufficient for a clinical decision (albeit a negative one, i.e. to exclude the antibiotic in question), in vitro susceptibility is necessary but not sufficient for a positive clinical decision (i.e. what antibiotic should be used). These same reasons have led to the increasing use and fortune of laboratory strategies aimed at the determination of resistance traits (from ß-lactamase tests to the molecular detection of specific resistance genes).
But while the relatively poor predictive value of susceptibility in vitro appears beyond dispute, does resistance in vitro, as currently defined, invariably predict therapeutic failure? In a recent clinical trial in Italy, most of the macrolide-resistant S. pyogenes strains isolated from patients who subsequently received macrolide therapy were nevertheless eradicated.19 Although a Pollyanna phenomenon (the fact that under particular conditions drugs with limited antibacterial activity may appear more efficacious than they really are25) could not be completely excluded because of the unavoidable absence, for ethical reasons, of an untreated control group, the fact that macrolides fared better than susceptibility studies had led to expect confirmed previous arguments for the in vivo activity of this class of drugs against some S. pyogenes strains categorized as resistant in vitro.26 In the context of streptococcal infections, the role of the level of antibiotic resistance in affecting the outcome of therapies with drugs showing suboptimal in vitro activity has already been suggested: (i) by the ability of penicillin to overcome in vitro penicillin resistance in pneumonia caused by Streptococcus pneumoniae strains with penicillin MICs up to 2 mg/L;2729 and (ii) by the poor correlation between the co-trimoxazole resistance of pneumococcal isolates and failures of therapy in children with pneumonia treated with co-trimoxazole.30
What do the above data mean? That a clinical isolate can be resistant in vitro to an antibiotic to which it responds in vivo? Obviously not. Genuine in vivo response does mean that the strain is not resistant; rather, it is the in vitro resistance that should be reconsidered. After all, resistance in vitro, like susceptibility, is a mere convention resting on established breakpoints. Over a decade ago, Sanders24 called the breakpoints magic to stress their crucial significance and at the same time the need to accept and apply them ready-made in routine diagnostic work: only organisms crossing this magic line come to be considered as resistant. In fact, although Walker & Thornsberry31 suggested that authors should only report MIC distribution rather than susceptibility categories, the reporting of antimicrobial susceptibility laboratory testing generally relies on the so-called SIR (susceptible, intermediate, resistant) system.
On the other hand, as Phillips22 recently pointed out, antibiotic resistance is not the definitive phenomenon that most of us treat it as. In particular, while the qualitative aspects of antibiotic resistance are increasingly well understood, our knowledge of its quantitative aspects continues to trail behind, and this is also true of attempts to establish antibiotic breakpoints with clinical validity. After initially being carefully determined based on microbiological, pharmacokinetic, pharmacodynamic and clinical data, the breakpoints should be re-evaluated periodically as changes in bacterial resistance, susceptibility testing methods or antibiotic formulations take place.32,33 Unfortunately, the entire matter shows a tendency to become more and more involved: the need to provide ever more reliable and predictive in vitro data has led, and will do so even more in the future, to an increasingly difficult and complicated interpretation of laboratory results. The multiplication of the relevant documents (on disc and MIC testing) issued by the NCCLS reflects this evolution well. In the early NCCLS documents (late 1970s to late 1980s), all antibiotic breakpoints were listed in a single, large but simple table.34,35 Between the late 1980s and the mid-1990s, additional tables began to appear listing separate antibiotic breakpoints for particular organisms: Haemophilus spp. (1988)36,37 was followed by Neisseria gonorrhoeae (1990),38,39 S. pneumoniae (1993)40,41 and streptococci including S. pneumoniae (1995).42 Starting from 1998, a new approach was adopted: the general table disappeared, to be replaced by distinct tables differentiated by groups of organisms. To the initial nine tables (Enterobacteriaceae, Pseudomonas aeruginosa and Acinetobacter spp., Staphylococcus spp., Enterococcus spp., Haemophilus spp., N. gonorrhoeae, S. pneumoniae, Streptococcus spp. other than S. pneumoniae, and Vibrio cholerae),43 a tenth table (Helicobacter pylori), limited to MIC testing, was added in 2000.44 Nor is this evolution likely to be over. As well as depending on the organism, antibiotic breakpoints are now beginning to be differentiated based on the site of infection: in the latest NCCLS document, different breakpoints of some ß-lactam antibiotics have been reported for pneumococci depending on their involvement in meningitis or other infections.45 The next steps are likely to bring further differentiation: for instance, the same MIC for the same type of organism might have a different meaning depending on the mechanism of resistance, e.g. target site modification or active efflux.
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