W. Stille

The Prognostic Value of the Antibiogram

The predictive value of the antibiogram is related to that of the MIC but is not completely identical. According to a common opinion, the effect of chemother- apy can be predicted by a simple comparison of the MIC and the concentration of the antibiotic at the site of action (Table 1). Such a simple balance-like scheme of chemo-

Table 1: Clinical effect as predicted by MICs in relation to the antibiotic level at the site of infection.

MIC > local level = no effect MIC = local level = doubtful MIC < local level = success

therapy has certain merits, e.g. in teaching students and nurses. This model of chemotherapy is valuable to a large extent for substances with a long half-life with homogene- ous distribution and constant activity under all conditions. This apparently applies to only a minority of the antimi- crobials available. With commonly used antibiotics - mainly [3-1actams and aminoglycosides - there are many arguments against both sides of this formula (Table 2).

Table 2: Factors which may modify the efficiency of anti- biotic therapy.

Therapeutic activity takes place: - in different tissue fluids; - with high and low inoculum; - often as mixed infection; - often in reduced oxygen tension; - after 2-6 h exposure to the antibiotic; - together with host defense mechanisms.

The real local levels of antibiotics are unknown to a large extent; they are not at all static. They increase and de- crease in one and the same infection. There are blood le- vels, tissue levels, intracellular levels and levels in body secretions such as bite and urine. I do not want to stress the critical aspects concerning local levels; there are even more arguments against the value of the MIC. There is the statistical argument. The MIC is the lowest concentration which inhibits the most resistant cell of the bacterial strain. There are further critical aspects of the MIC. The MIC is relatively inaccurate; it is measured in broth with a stand- ardized inoculum. In clinical chemotherapy, however, in-

fections with only very few bacteria and other infections with many bacteria may be present. 30% of all infections are mixed infections; the rules of monoinfections do not apply in polymicrobial infections. Sensitivity tests are done in a normal atmosphere, but many infections take place under anaerobic conditions or under reduced oxy- gen tension. MICs are performed with a constant level of antibiotics, but in routine testing for MICs nobody evalu- ates whether enzymatic inactivation takes place during the procedure. If we say minima] inhibitory concentra- tion, we do not know how much antibiotic we really have at the end of the test. Disc sensitivity tests as a simplified form of the MIC add additional irrational points to the antibiogram. Generally, antibiograms are not very accurate due to inoculum ef- fects. In disc tests with cefotaxime and Enterobacter cloa- cae, many zone diameters can be found using different in- ocula. Disc tests should be performed with a standardized inoculum. If there is an error in the inoculum, another zone size will be achieved. There are some antibiotics where one can get any MIC one wants by using different inocula (Figure 1).

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0.125 0.25 0.5 1 7 t, 0 10 32 0t, mg/t

Figure 1' Cumulative distribution of inoculum-dependent MICs of fosfomycin for seven Escherichia cotistrains (in- oculum 10-10 8 cfu/ml).

In substances like cefsulodin, the MIC distribution is very close together (Figure 2); this is the way it should be. An optimal antibiotic always shows the same activity with a low or high inoculum.

Summary and literature references were not received for this manu- script. Prof. Dr. W. Stille, Klinikum der Universit~it, Zentrum Inhere Medizin, Theodor-Stern-Kai 7, D-6000 Frankfurt 70.

S 66 Infection 11 (1983) Suppl. 2

W. $tille: Prognostic Value of the Antibiogram

100 %

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Figure 2: Min imum inhibi tory concentrat ions of cefsulodin against Pseudomonas aeruginosa(n = 20). Inoculum: 103, 10 s and 10 r cfu/ml. (Agar dif fusion test).

E. c0li

6363 Cephalexin

=O.t,

=0.2

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PLOT frequency = 30 rnin

Figure 4: Partial activity of cephalexin against Escherichia coil strain 6363.

MICs are biological parameters. They cannot be compar- ed with exact physical or chemical values. An MIC of 5 mg/l does not mean that 4.9 mg/1 are completely in- active. From the point of view of breakpoint, a border- line concentration is not possible. If at all, it is a break "range", not a point. There is a wide range of partial activity below the MIC. This can be analyzed by registering growth curves (Figure 3). Partial activity may be of very different types: I. slight substitution with a slight delay of growth; 2. a real de- crease in velocity of growth; 3. paradoxical effects usually due to filament formation and 4. inactivation curves. Some antibiotics have a small range of partial activity; others have quite a marked range of partial activity. As an example, cephalexin has a small range of partial ac- tivity (Figure 4). Typical inactivation curves can be found with cefaclor (Fi- gure 5). To see the problem in another way, the activity of cefaclor is much higher than the activity measured by the MIC. The range of partial activity is important for the ex- tent of general activity. An analysis of partial activity may E c01i give us some important information concerning a new an- tibiotic. The range of partial activity can be connected 63153 with intermittent chemotherapy. Cefaclor

The activity of an antibiotic in decreasing concentrations may act as a model of intermittent chemotherapy (Figure

=G

=2

0 =1

Figure 3: Four main types of subinhib i tory growth curves.

6). We start the system with twice the MIC. Below the MIC, i. e. with 50-25% of the MIC, there is regrowth with Streptococcus faecalis, but there is no immediate re- growth with Proteus mirabilis. There is a postantibiotic ef- fect below the partial activity, an aspect which is not con- sistent with the so-called chemotherapeutic formula. If physicians draw their conclusions from animal experi- ments, it is always justified to argue about the remarkable difference between men and mice. Nevertheless, it is a generally accepted procedure to draw conclusions from activity in peptone broth to the activity in the human bo- dy. One thing is absolutely clear, however: the human bo- dy is free of peptone and similar amino acid solutions. It is not very difficult to evaluate the MICs of different anti- biotics in certain body fluids. Kanamycin and gentamicin are tested under the same conditions with a Klebsiella strain (Figure 7). A complete loss of activity can be found when urine agar is used in- stead of DST medium. There is no activity in solidified urine. When MICs for aminoglycosides were determined

1.25 mg 2.5 mg

/ . . . . . . . . _ ~ 5 mg/[

P10T frequency 30 rnln Figure 5: Partial activity of cefaclor against E$cherichia coli strain 6363.

Infection 11 (1983) Suppl: 2 S 67

W. StOle: Prognostic Value of the Antibiogram

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Figure 6: Antibiotic activity in de- creasing concentrations as a model of intermittent chemotherapy.

in urine, there was a remarkable loss of activity. The dif- ferences between activity in the test medium and activity in body fluids can be measured as the "relative activity". With tetracycline, the relative activity in different human body fluids, such as bile, amniotic fluid, plasma water and urine, indicates at least a remarkable loss of activity against E. coli, but not against Proteus (Table 3). There can be differences with different species and there are re- markable differences between different types of body fluids. A substance with good antibacterial activity such as ampi- cillin loses much of its activity if you test the kill-rate in amniotic fluid, bile or plasma water (Figure 8). Antibiotics like fosfomycin may lose their not very mark- ed activity completely. Tests of the bacterial activity in

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Figure 7: Disc test for kanamycin and gentamicin against a Klebsiella strain on urine agar (left) and on DST medium (right).

bile or urine show no antibacterial activity at all (Figure 9). The interactions between body fluids and test medium cannot be explained by differences in pH alone. The poor activity of tetracyclines in bile may explain the poor clinical results of treatment of biliary tract infections with these antibiotics. This apparently has a clinical equival6nt in the poor elimi- nation of sensitive E. coli from the bile during treatment of E. coli infections with tetracycline. With doxycycline, the number of bacteria in the bile during biliary drainage stays at the same level. With mezlocillin, eradication of E. coli from the bile can be achieved within 48 hours (Figure 10). There are many other examples showing the supe- riority of betalactams over tetracyclines.

1000 %

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F i g u r e 8: K i n e t i c s o f t he b a c t e r i c i d a l ac t i v i t y o f a m p i c i l l i n (10 mg/I) against Escherichia colistrain 6363 in body fluids and in bouillon. Inoculum: - 105 cfu; method: membrane filtration•

S (18 Infection 11 (1983) Suppl, 2

...... Bi te

J Urine

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Figure 9: Kinetics of the bactericidal activity of fosfomycin (100 mg/I) against Pseudomonas aeruginosa strain 4833. MICs in bile > 512 mg/t; in urine 32 mg/I; in Mueller-Hinton broth 16 mg/l.

In certain bacteria, there can be true clinical resistance despite in vitro sensitivity. This may be present with oxa- cillin-resistant or penicillin-tolerant staphylococci and with ampicillin-resistant Haemophilus influenzae. Oxacil- lin resistance in Staphylococcus aureus is heterogeneous and may mimic sensitivity in normal disc tests despite cli- nical failure. The problem can be solved by testing in hy- pertonic media, or by testing at a lower temperature. A comparable problem can be found in penicillin-tolerant staphylococci. Ampicillin-resistant 14. influenzae may be sensitive in disc tests, but clinical treatment is ineffective due to the strong [5-1actamase. Another aspect of chemotherapy must be stressed. MICs are generally unreliable for slow growing bacteria, for many bacteria with degradating enzymes (mainly Bactero- ides fragilis), for other anaerobic bacteria and for fastidi- ous agents. The concept of directed chemotherapy acts

10 7

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Figure 10: Number of Escherichia coliorganisms (sensitive in vitro) in bile during treatment with doxycycline and me- zlocillin. DOXY = doxycycline; MEZLO = mezlocillin.

W. Stille: Prognostic Value of the Antibiogram

well in rapidly growing normal pathogens, but it is of no value in slow growing strains. If one looks for the practi- cal evaluation of MICs and antibiograms, it is a field where errors and even falsifications may be present. With some antibiotics it is possible to modify the conditions of the MIC in order to achieve optimal activity in vitro. For example, fosfomycin will only show good activity against resistant bacteria in a medium with a high content of glu- cose-6-phosphate - a condition which is certainly not pre- sent in the human body. The activity of fosfomycin in hu- man body fluids is much lower than its activity in normal peptone broth. Under the right conditions with the right trick and the right additive, a low-quality antibiotic may mimic a highly active substance. Additives to a medium are only justified to paralyse antagonists to the antibiotic, but not to achieve an optimal in vitro activity.

Table 3: Range of relative activi ty of tetracycl ine against Escherichia coli, Proteus mirabilis and Klebsiella pneumo- niae in di f ferent body fluids.

/ Plasma water 1/128--1/32 1-2 1/128-1/64 4 ]

t Bile lho24-itc~ ifs-dt4-1h ift024-1112s 6 A m n i o t i c f l u i d 1/1024-1/256,-l/4 l / 4 - s m 112048-l/512--1 5 ;'

Urine Vs~/4-l/4 I/8J/8-1/4 VzJ/2-1 6

The contrary is also possible. Some antibiotics have good clinical activity and poor in vitro activity. This is the case in usual testing with co-trimoxazole and to a certain ex- tent with chloramphenicol. The activity of an optimal an- tibiotic should always be in the same range under differ- ent conditions. A substance for which any MIC can be achieved by only varying the conditions or the inoculum slightly, or a substance which loses its activity in several body fluids is at least a doubtful agent. Other information can be obtained from an antibiogram. The antibiogram provides certain taxonomic information. The resistance of a strain may give some information on the species of bacteria, e. g. the lincomycin resistance in S. faecalis and the polymyxin resistance in Proteus species and Serratia marcescens. A strain with isolated resistance to cefoxitin is most likely a strain of Citrobacter (Table 4). The antibiogram can give us additional and more general information. It may help in the identification of strains and it can give us information concerning group resist-,~ ance. In E. coli, resistance to ampicillin is a sign of active [5-1actamase production. Further treatment should only consist of substances which are completely stable to [Mac-

Table 4: Taxonomic information from the antibiogram.

Lincomycin Polymyxin

Cefoxitin

= Streptococcus faecalis = Pro t eus sp.

Serratia marcescens

= Ci t robac ter

= resistance

Infection 11 (1983) Suppl. 2 S 69

W. Stille: Prognostic Value of the Antibiogram

tamases. Despite the great merits of antibiograms, espe- cially in UTI, there is no doubt that antibiograms are abused in practical medicine, at least in Germany. Spu- tum cultures of innocent bacteria like Streptococcus viri- dans, Neisseria, E. coil and Staphylococcus epidermidis, are frequently tested with antibiograms: a lack of infor- mation as well as commercial interests of bacteriologists and clinical pathologists could be the cause of such fancy antibiograms. I do not wish to further stress the weak aspects of MICs, but I do want to draw your attention to the clinical corre- lation of sensitivity in vitro and clinical success. Without a doubt, there are many instances where directed chemo- therapy acts according to the chemotherapeutic formula: the strain is sensitive, there is a sufficient level at the site of action and success is inevitable. Whether the real con- ditions are as simple as it seems is another problem. In modern chemotherapy, failures are more interesting than successes. There are remarkable exceptions concerning the effect of chemotherapy. There may be activity in vitro, but no clinical effect (Table 5). A classical example is the treatment of typhoid fever.

Table 5: Examples of clinical failure despite in vitrosensi- tivity of the isolate.

Typhoid fever Cephalosporins Aminoglycosides Tetracyctines

Streptococcus faecalis Penicillin G endocarditis Ampicillin

(monotherapy)

Severe local infections Polymyxins by enterobacteria Aminoglycosides

Many antibiotics (cephalosporins, aminoglycosides and tetracyclines) are sensitive to Salmonella typhi in vitro, but treatment is not effective. Another example, mono- therapy with penicillin G or ampicillin, leads to insuffi- cient success in enterococcal endocarditis. Recovery is usually not achieved. This failure of monotherapy with penicillin is apparently due to the insufficient bacterial ac- tivity. High concentrations of these antibiotics lead to pa- radoxical effects in S. faecalis, the so-called Eagle effect. Only in combination with gentamicin, which is nearly in- active as a single agent, is it possible to treat enterococcal endocarditis effectively. Clinical failures can be related to poor penetration of the antibiotics into the site of action. An example is the poor activity of polymyxins and, to a lesser extent, aminoglyco- sides in severe tissue infections caused by enterobacteria and Pseudomonas. The opposite is also possible. In some instances, clinical effects can be seen despite resistance in vitro (Table 6). This may be found in UTI or superficial wounds. Break- points in sensitivity testing are oriented to a medium blood level, but local levels may be 10, 100 or 1000 times higher than this value. Another example is once again the

Table 6: Clinical success of antibiotic treatment with drugs ineffective in vitro.

Urinary tract infections, Effect by high superficial wounds local levels

Gentamicin in enterococcal Combination effect endocarditis

combination effect of gentamicin in enterococcal endo- carditis. In some instances, the differences between in vitro and in vivo activity are not so marked. The high in vitro activity of cefotaxime would suggest treatment with 150300 mg daily. Such doses are apparently inactive despite the high in vitro activity. The clinical dose of cefotaxime cannot be reduced below the dosage of conventional cephalospo- rins. The contrary is also possible. The clinical efficacy can be better than the MICs predict. This is mainly true with chloramphenicol. Good clinical effects are obtained despite MICs which are just barely in the range or even below the blood levels. The explanation for this is, in our opinion, an antagonism of phenylalanine in the medium. The real activity of chloramphenicol is up to eight times higher than the MIC. We have comparable data concern- ing the activity of carbenicillin against Pseudomonas. The real activity seems to be higher than the MIC. The activity in body fluids is higher than the activity in broth which ap- parently contains an antagonist to carbenicillin. It should be stressed that chemotherapy directed accord- ing to this model is only justified in monobacterial infec- tions in which the pathogen is known. This only applies to a minority of infections. In general, the other types of chemotherapy are more important for the practical treat- ment of infections. We should consider the general value of antibiograms and MICs more than their individual value. The MIC and the antibiogram are often regarded by the clinician as a positive order to use a special antibiotic. This is wrong! It is not an order to use one or several anti- biotics. The selection of the optimal antibiotic for the ac- tual infection must be based on several additional factors like site of infection, pharmacology, prior intolerance and underlying diseases (Table 7). The art of medicine today thus consists to a large extent of the selection of the opti- mal antibiotic from a remarkable number of agents.

Table 7: important clinical and pharmacological aspects of antibiotic therapy.

The antibiogram gives information about resistance of certain antibiotics. For selection of the optimal antibiotic for the actual infection, several additional factors have to be considered:

Site of infection Pharmacology of the antibiotic Prior intolerance Prior antibiotics Concomitant medication Underlying diseases

S '70 Infection 1t (1983) Suppl. 2