Confirm need: identify target; keys to appropriate antimicrobial use

Q: Please review interpretation of bacterial culture and sensitivity testing.

A: Dr. Dawn M. Boothe at the 2005 American College of Veterinary Internal Medicine (ACVIM) Forum in Baltimore gave a lecture on appropriate use of antimicrobial drugs. Some relevant points in this lecture are provided below.

Important considerations for selection of an antimicrobial are, in order of priority: 1) confirming the need; 2) identifying target organisms; and 3) identifying the target site.

Additional considerations should include identifying mechanisms of resistance, the likelihood of achieving bactericidal concentrations at the site, and time — versus concentration — dependency as it relates to convenience. Cost should be the last consideration that influences selection.

Antimicrobial selection should strive for a spectrum that is as narrow as possible, thus avoiding the sequelae of drug use on the normal flora and unnecessary selection pressure. For complicated infections, identification should be based on culture and sensitivity; this information provides a basis for designing a dosing regimen most appropriate for the animal.

Basing antimicrobial selection on culture and sensitivity information does not guarantee success, just as failing to use culture and sensitivity as a basis for selection (or selecting a drug characterized by "R" on the information) does not guarantee failure. The "90-60 rule" implies that approximately 90 percent of infections treated based on culture and sensitivity are likely to respond if an "S" drug is selected; yet, up to 60 percent will respond even if an "R" drug is selected. The most likely situation where the latter is true is if the infection is at a site in which the drug is prescribed at a much higher dosage than that achieved in the test tube (i.e., much higher than the minimum inhibitory concentration (MIC).

Although culture and sensitivity information can be a powerful tool to guide selection, it nonetheless is an in vitro test applied to in vivo conditions; over-reliance on the information can contribute to therapeutic failure. Anaerobic infections are particularly problematic. Obligate anaerobes are exquisitely sensitive to increased oxygen tension and will not survive if exposed to oxygen. No growth may be mistakenly interpreted as lack of anaerobic infection. Many organisms are facultative anaerobes, capable of growth in anaerobic environments. Aerobic cultures may yield their growth, but the anaerobic environment in the animal may limit response to antimicrobials (particularly aminoglycosides). Organisms without cell walls (Mycoplasma, L-forms, etc.) and others difficult to culture, or slow-growing organisms (anaerobes, selected Gram positives, Nocardia, atypical Mycobacteria and others) and culture and sensitivity information may not include MIC.

Just as absence of growth does not indicate absence of infection, isolation of an organism is not necessarily evidence of infection, nor even if infection is present, does the isolated organism represent the infecting organism. Clearly, culture of an organism from a tissue that is normally sterile indicates infection. However, discriminating between normal and infecting flora can be difficult. Pure culture and large number of organisms are indicators of infection.

The culture and sensitivity procedures themselves are fraught with potential errors. For practices that provide in-house susceptibility testing, care should be taken to follow guidelines established and published by (or comparable to) the Clinical and Laboratory Standards Institute (CLASI) or comparable federal agency. Materials, including interpretive standards, should be validated by the appropriate agency. Minor changes in pH, temperature, humidity, etc. can affect results profoundly. Personnel should be trained specifically in culture techniques, and hospitals that provide this service should maintain well-designed and adequately collected quality control data to validate their procedures.

Pitfalls of susceptibility testing also reflect the drugs selected for testing. Not all companies are interested in establishing interpretive criteria and as such, not all drugs are available for testing. Because automated systems cannot accommodate, and laboratories (nor owners) cannot afford, testing all potential drugs used to treat an infection, one drug often is tested as a model for other drugs in the class (i.e., cephalothin for the first-generation cephalosporins; enrofloxacin for the fluorinated quinolones). For some classes of drugs, cross-reactivity can be similar within the class [for example, an organism that is R to one fluorinated quinolone (including ciprofloxacin) is likely to be R to all]. However, the same is not true for others. Amikacin is often more effective than gentamicin (hence both are often on a report). Cephalothin serves as a model for all first-generation cephalosporins, but it underestimates efficacy of cefazolin toward Gram-negative organisms. The spectrum of third- and fourth-generation cephalosporins is too variable to allow one to predict the susceptibility of others and as such, multiple drugs are likely to be included. Culture and susceptibility techniques may not accurately reflect resistance that has developed in the infecting organism to drug to which the organism is generally susceptible. These cephalosporins also offer another example of concern: they are susceptible to extended spectrum beta-lactamase that will be produced in vivo but not in vitro and despite an "S" designation, therapeutic failure may occur. Ceftazidime is often used as a test for the presence of extended spectrum beta-lactamase; imipenem and clavulanic acid generally are not susceptible to these enzymes. Laboratories currently are generating methods intended to detect this level of resistance.

Interpreting culture and sensitivity (pharmacodynamic) information is most helpful when considered in the context of the animal. The first step should focus on comparing what is needed — the MIC of the isolate for the drug of interest — to what is achieved. Even if samples are properly collected and cultured, culture and sensitivity information is inherently deficient because in vitro methods cannot mimic in vivo conditions. For example, the isolate is exposed to the constant conditions, including drug concentrations throughout the in vitro incubation period; in vitro methods cannot take into account host factors that detract from efficacy. Among the most problematic concerns is interpretation of MIC. The breakpoint MIC (MICBP) is based, in part, on peak plasma drug concentrations (Cmax) which, ideally, is determined in the species of interest and tested against pathogens infecting the targeted animals.

Many drugs used by veterinarians are approved for use in humans. Although interpretive MIC data has been determined by CLASI for some drugs in animals, many have not and interpretation may be inappropriate. Ciprofloxacin is an excellent example. Its oral bioavailability in dogs is 30 percent to 40 percent of that in humans, and despite its increased potency compared to enrofloxacin toward Gram-negative organisms, its potential efficacy (MICBP) is equivalent to or less for many organisms. Susceptibility data also does not take into account active metabolites, again exemplified by enrofloxacin, which is metabolized to ciprofloxacin: both Cmax and area under the curve (AUC) of bioactivity of enrofloxacin may increase up to 50 percent or more by ciprofloxacin; as such, culture and sensitivity information might underestimate efficacy.

MICBP generally are based on the highest labeled dose, but higher doses might be safely administered for many antibiotics. If recommended doses change, the manufacturer should provide CLASI with updated pharmacokinetic information so that interpretive criteria may change accordingly, and automated systems should incorporate those changes in their methods. Again, enrofloxacin offers an example. Originally approved at 2.5 mg/kg, 1 microgram/ml (< 1= S; > 2 =R) was the MICBP; the current dose is up to 20 mg/kg and new interpretive criteria identify > 4 micrograms/ml = R.

The term "bactericidal" is somewhat abused. Veterinarians will reach for a drug that is "cidal" rather than "static", assuming that the ability of the drug to kill rather than simply inhibit organisms will increase the risk of therapeutic efficacy. This may be true, but only if concentrations of the drug achieved at the site of infection are sufficient to kill the microbe. The term "bactericidal" is an in vitro definition and is based on killing rates (e.g., 99.9 percent reduction in bacterial inoculum within a 24-hour period of exposure) as well as the proximity of the minimum bactericidal concentration (MBC) of a drug to the MIC. The MBC is determined following tube dilution procedures; tubes with no observable growth are inoculated on agar gel. If no organism grows on the agar, the organisms were killed in the test tube. The tube with the lowest concentration of drug that yields no growth on the agar gel contains the MBC of drug. For drugs considered bactericidal, the MBC is within one tube dilution of the MIC, meaning, the organisms were not simply inhibited, but rather, were killed.

For "bacteriostatic" drugs, growth on the agar plate will occur for several tube dilutions above the MIC, indicating that organisms were not killed. However, bacteriostatic drugs are capable of killing [e.g., some organisms are exquisitely sensitive to the effects of selected drugs; some static drugs are accumulated to concentrations that are likely to be cidal (e.g., macrolides and lincosamides in phagocytes; urine concentration)]. However, killing concentrations are generally more likely achieved for a cidal drug compared to a static concentration. On the other hand, a cidal drug may not kill if concentrations are not sufficient, or if conditions preclude its actions (e.g., slow growth in an anaerobic environment; combination with growth inhibitors). Thus, cidal effects will occur only if adequate concentrations (i.e., MIC/MBC) are achieved at the site. The bactericidal nature of a drug often reflects its mechanism of action.

Drugs that target ribosomes (e.g., tetracyclines, macrolides, lincosamides, chloramphenicol) often simply inhibit the growth of the organism, and, because a much higher drug concentration is necessary to kill the organism, in vitro, the MIC is distant from the MBC. Clinically, host defenses must eradicate the infection following treatment with these drugs unless exceptionally high concentrations (i.e., the MBC) of these drugs are achieved in tissues. An exception is made for aminoglycosides, whose ribosomal inhibition is so effective that the organism dies.

Drugs that target cell walls (beta lactams including penicillins and cephalosporins; vancomycin), cell membranes (bacitracin, polymixin and colistin), and DNA (enrofloxacin, metronidazole) and RNA (rifampin) are defined in vitro as bactericidal. Combinations of static drugs can often result in "cidal" actions. For example, sulfonamides (which target folic acid synthesis) are "static", but when used in combination with diaminopyrimidines (e.g., trimethoprim), the combination is defined in vitro as cidal.

Attaining bactericidal concentrations of an antimicrobial is critical for those infections for which host killing is likely to be impaired. These include, but are not limited to, infections in immune-compromised animals (e.g., viral infection like parvovirus, panleukopenia, FIV, FeLV), animals receiving glucocorticoids or in systems characterized by derangements in local immunity (i.e., CNS infection for which a marked inflammatory response can be life threatening; osteomyelitis; peritonitis, bacteremia/sepsis, many chronic infections).

Dr. Johnny Hoskins is owner of DocuTech Services. He is a diplomate of the American College of Veterinary Internal Medicine with specialities in small animal pediatrics. He can be reached at (225) 955-3252, fax: (214) 242-2200 jdhoskins@mindspring.com