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Antimicrobial Agents and Chemotherapy, August 1998, p. 1973-1979, Vol. 42, No. 8
0066-4804/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
In Vitro Activities of Oral 
-Lactams at Concentrations
Achieved in Humans against Penicillin-Susceptible and
-Resistant Pneumococci and Potential to Select Resistance
Christine E.
Thorburn,1,*
Sarah J.
Knott,1 and
David I.
Edwards2
SmithKline Beecham Pharmaceuticals, Brockham
Park, Betchworth, Surrey RH3 7AJ,1 and
Chemotherapy Research Unit, University of East London,
London E15 4LZ,2 United Kingdom
Received 31 October 1997/Returned for modification 9 February
1998/Accepted 31 May 1998
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ABSTRACT |
The 
-lactam susceptibilities of 65 strains of
Streptococcus pneumoniae for which penicillin MICs covered
a broad range were assessed. The order of potency was amoxicillin (AMX) = amoxicillin-clavulanate (AMC) > penicillin G > cefpodoxime (CPO) > cefuroxime (CXM) > cefprozil > cefaclor > loracarbef > cefixime. No decrease in
susceptibility was seen following repeated subculture of two
penicillin-susceptible strains of S. pneumoniae in AMX, AMC, cefaclor, or loracarbef, whereas repeated exposure to CPO and CXM resulted in 4- to 32-fold decreases in susceptibility for both strains. When one of these strains
was exposed to concentrations of CPO, CXM, AMX, and AMC achieved
in the serum of humans following the administration of an oral dose,
all agents were rapidly bactericidal, with no decrease in
susceptibility up to 72 h. This was consistent with
antibiotic concentrations exceeding the MICs for 100% of the
dosing interval. For a penicillin-resistant strain, MICs were exceeded
for 29% of the 12-h dosing interval for 500 mg of AMX, 42% of the
interval for AMC with 875 mg of AMX and 125 mg of
clavulanate (875/125 mg of AMC) 21% of the interval for
500 mg of CXM, and 0% of the interval for 200 mg of CPO. Consequently,
only 875/125 mg of AMC produced a sustained bactericidal effect. A
four- to eightfold reduction in susceptibility to CPO and CXM and
cross-resistance with cefotaxime, but not penicillin or AMC, were
selected following exposure to simulated serum CPO and CXM
concentrations. In addition, AMX and AMC were the only agents which
consistently produced a >99% reduction in bacterial numbers in
time-kill studies using concentrations of antibiotic achieved in middle
ear fluid for all three strains of penicillin-resistant S. pneumoniae tested.
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INTRODUCTION |
The incidence of penicillin
resistance is increasing among clinical isolates of Streptococcus
pneumoniae. Penicillin-resistant pneumococci have been present at
high frequencies (44 to 59%) in South Africa (23), Hungary
(28), and Spain (13) for some time, but rates of
>40% have now also been reported in France, Argentina, Uruguay,
Mexico, Israel, Saudi Arabia, Nigeria, Kenya, Japan, and Korea
(24). Penicillin-resistant pneumococci are often also
resistant to a number of other classes of antibiotics, such as
tetracyclines, chloramphenicol, and
trimethoprim-sulfamethoxazole (1, 13), making the choice
of therapy difficult. The increasing incidence of macrolide
resistance (13, 15), which confers a high level of
resistance to all currently available agents in that
class, also means that agents such as erythromycin, clarithromycin, and
azithromycin will frequently be less effective. It is hoped that the
development of quinolone antibiotics with improved activity against
gram-positive bacteria will overcome doubts as to the usefulness of
these agents in treating pneumococcal infections (27). The
use of quinolones is contraindicated in children and infants, however,
and many have further toxicity problems which restrict their use. These
agents also have a higher potential than, for example, -lactams and
macrolides to select for mutational resistance, and there are
documented cases of clinical failures that can be directly related to
the emergence of bacterial resistance following quinolone therapy
(31, 32).
Resistance to other antibiotic classes means that orally administered
-lactam antibiotics may still be the first choice for empiric
therapy of community-acquired respiratory tract infections, even in
countries with a high incidence of penicillin-resistant pneumococci,
because the variable or nonabsolute nature of this resistance means
that some -lactam antibiotics remain efficacious (20). It
is therefore of great importance to choose the most potent -lactam
antibiotic against the key respiratory tract pathogens (S. pneumoniae, Haemophilus
influenzae, and Moraxella catarrhalis), to
use it at a dose high enough to be effective against
penicillin-resistant strains of S. pneumoniae, and to
choose the compound least likely to select a higher level of -lactam
resistance.
In the studies described here, the in vitro potencies of a number
of orally administered -lactam antibiotics against
S. pneumoniae were compared, as were their potential to
select for resistance. The effectiveness of the antibiotic
concentrations achieved in middle ear fluid (MEF) against
penicillin-resistant pneumococci was examined, and for the most
potent agents, the concentration-time profiles achieved in the serum of
humans following the administration of a conventional oral dose were
simulated in an in vitro pharmacodynamic model. This was used to assess
bactericidal activities and the potential to select for resistance
among penicillin-susceptible and penicillin-resistant pneumococci.
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MATERIALS AND METHODS |
Compounds.
Amoxicillin trihydrate and potassium
clavulanate were supplied as laboratory reference standards
by SmithKline Beecham Pharmaceuticals (Worthing, United Kingdom).
Cefuroxime was used as the commercially available injectable
preparation (Zinacef; Glaxo); cefaclor (Distaclor; Dista Labs)
and cefixime (Cefspan; Fujisawa) were extracted from the commercially
available capsules; and cefpodoxime (Roussel), cefprozil (Bristol Myers
Squibb), and loracarbef (Eli Lilly & Co.) were all kindly supplied as
soluble powders by the manufacturers.
Bacterial strains.
The MICs for a number of clinical
isolates of S. pneumoniae from France, Hungary, South
Africa, and the United Kingdom with a range of susceptibilities to
penicillin were determined by the agar dilution method. A
penicillin-susceptible type strain, S. pneumoniae ATCC
6303, and a penicillin-susceptible clinical isolate from Hungary,
S. pneumoniae 5303, were used in the serial passage experiments. Three penicillin-resistant strains of S. pneumoniae (one from South Africa [strain N1387] and two
S. pneumoniae strains kindly supplied by Dr. Ridgeway,
Prescot, United Kingdom [strains R1 and R2]) which had different
patterns of susceptibility to the test -lactam antibiotics (Table
1) were chosen for the time-kill studies.
A type strain, S. pneumoniae ATCC 6303, and a
penicillin-resistant clinical isolate, S. pneumoniae
1320b, supplied by Jenny Dahl Knudsen (Copenhagen, Denmark), were used
in studies with the in vitro pharmacodynamic model. S. pneumoniae R6, a susceptible control strain, and S. pneumoniae R61a2x, a laboratory-derived variant of R6 with
alterations in penicillin-binding proteins (PBPs) 1a and 2x
(6), were both kindly provided by Chris Dowson (Sussex University, Brighton, United Kingdom).
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TABLE 1.
-Lactam MICs for strains of S. pneumoniae used in time-kill studies with concentrations of
antibiotics achievable in MEF
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MIC determinations.
Serial twofold dilutions of antibiotic
were prepared in Mueller-Hinton agar (BBL) supplemented with 5%
(vol/vol) sterile defibrinated horse blood. The agar was inoculated
with each test organism at 105 CFU/spot, and the plates
were incubated for 18 to 24 h at 37°C. The MIC was the lowest
concentration of antibiotic that completely inhibited visible bacterial
growth.
Serial passage experiments.
A series of twofold dilutions of
twice the final required concentrations of antibiotic were prepared in
1-ml volumes of Todd-Hewitt broth (Oxoid) supplemented with 5%
(vol/vol) heat-inactivated horse serum. Overnight broth cultures of the
test organisms were diluted to the turbidity of a 0.5 McFarland barium
sulfate standard, and a further 10-fold dilution was made in
Todd-Hewitt broth plus 5% serum. Each antibiotic concentration was
inoculated with 1 ml of the test culture to give approximately 5 × 106 CFU/ml. The MIC of each antibiotic was determined
following 24 h of incubation at 37°C, and the culture containing
the highest antibiotic concentration allowing visible growth was
adjusted to the turbidity of a 0.5 McFarland standard. Following a
10-fold dilution, this culture was used to inoculate a fresh series of antibiotic dilutions, as described above. This process was repeated for
5 days. The final MICs were noted, and the end-of-passage isolates were
tested for stability of resistance and cross-resistance with other
agents by a conventional agar dilution MIC determination procedure as
described above.
Time-kill studies.
Solutions containing the antibiotics at
the peak concentrations reported to be achieved in the MEF of children
(Table 1) were prepared in 20-ml volumes of Todd-Hewitt broth (Oxoid)
supplemented with 5% (vol/vol) heat-inactivated horse serum. The media
were inoculated to give 105 to 106 CFU/ml and
were incubated at 37°C on an orbital shaker. Samples were taken for
assessment of the numbers of viable bacteria at 0, 1, 3, 5, 7, and
9 h. Serial 10-fold dilutions of the samples were prepared in
Todd-Hewitt broth, and four dilutions were plated in triplicate onto
nutrient agar (Lab M) supplemented with 5% (vol/vol) sterile horse
blood and 0.4% -lactamase (Penase; Difco) to inactivate the
antibiotic that was carried over. The numbers of CFU were determined
following 24 h of incubation at 37°C, with the limit of
detection being 1.67 × 101 CFU/ml.
In vitro pharmacodynamic model.
The open, one-compartment
model used was based on the biexponential model originally described by
Grasso et al. (16) in 1978, and is shown in Fig.
1. The flow rate of the pump and the volumes in the flasks were set to simulate the elimination rate of the
antibiotics with the shortest half-life (t1/2;
i.e., t1/2 = 1 h for amoxicillin,
cefuroxime, and clavulanate), whereas cefpodoxime (t1/2 = 2 h) was added at regular intervals
to simulate its slower elimination from humans. The dilution rate of
the bacterial cultures in the open system was therefore the same for
all of the test antibiotics. Repeated doses of antibiotic were
administered automatically every 12 h with a pump (Watson Marlow)
interfaced to a microcomputer (Commodore). Mueller-Hinton broth (Difco)
supplemented with 5% (vol/vol) sterile, heat-treated horse serum was
used. Samples were removed from the culture flask at regular time
points for determination of the concentration of antibiotic and the
number of viable bacteria present. Viable bacterial counts were
determined as described above for the time-kill studies, and the
antibiotic concentrations were assayed microbiologically. Colonies from
the viable count plates were tested for -lactam susceptibility by determination of the MIC by the agar dilution method to detect any
selection of resistance.
Microbiological assays.
Amoxicillin was assayed with a
commercially available Bacillus subtilis NCTC 6633 spore
suspension (Difco) in nutrient agar (Lab M), and
clavulanate was assayed with Klebsiella
pneumoniae NCTC 11228 in nutrient agar supplemented with 60 µg
of benzylpenicillin/ml (19). Cefuroxime and cefpodoxime were
assayed with Escherichia coli ESS (an extrasensitive
permeability mutant) in nutrient agar.
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RESULTS |
MIC determinations.
The penicillins amoxicillin (alone and in
the presence of clavulanate) and penicillin G were the most
active -lactams against penicillin-susceptible (Pens)
and penicillin-intermediate (Peni) pneumococci.
Amoxicillin was also the most potent agent against penicillin-resistant (Penr) S. pneumoniae, whereas cefpodoxime was more active than
penicillin G against some strains (Table
2). The most active cephalosporins tested
were cefpodoxime and cefuroxime, which are both available as oral ester
formulations. Cefprozil was two- to fourfold less active than
cefuroxime overall, and cefaclor was less active than cefprozil, although cefaclor was slightly more active
than loracarbef. Interestingly, cefaclor and loracarbef
were more active than cefixime against some strains of
S. pneumoniae but were less active against others.
Clavulanate was the least active compound tested and is not available
for clinical use alone but is available as a
-lactamase
inhibitor in
combination with amoxicillin. It was included as
a reference for the
amoxicillin-clavulanate studies, and because
it binds
selectively to PBP 3 in
S. pneumoniae (
33),
it was
of interest to see whether the activity of
clavulanate was affected
in the same way as the activities
of the other agents by altered
PBPs in Pen
r pneumococci. In
fact, the MIC at which 90% of strains are inhibited
(MIC
90) for clavulanate increased 64-fold
between the Pen
s and Pen
r strains, which
was similar to the effect seen for the other agents
tested (32- to
128-fold increases) except penicillin G, the MIC
90 of
which increased 256-fold, and cefixime, the MIC
90 of
which
increased only 8-fold.
Serial passage experiments.
No decrease in susceptibility was
seen following exposure of S. pneumoniae ATCC 6303 and
S. pneumoniae 5303 to amoxicillin, amoxicillin-clavulanate, cefaclor, or loracarbef
for five subcultures. Following exposure to cefpodoxime, however,
S. pneumoniae ATCC 6303 showed a fourfold decrease in
susceptibility to this compound (Table
3), whereas the culture passaged in
cefuroxime not only was fourfold less susceptible to cefuroxime but was
also fourfold less susceptible to cefpodoxime and cefixime.
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TABLE 3.
Agar dilution MICs of test -lactams for isolates of
S. pneumoniae ATCC 6303 and S. pneumoniae 5303 following five serial passages through
increasing antibiotic concentrations
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Following repeated subculture in the presence of cefixime,
S. pneumoniae 5303 became fourfold less susceptible to
amoxicillin-clavulanate,
although the
susceptibility to the cephalosporins was unchanged.
This strain became
fourfold less susceptible to cefpodoxime, cefixime,
and loracarbef and
eightfold less susceptible to cefuroxime following
5 days of exposure
to increasing concentrations of cefpodoxime.
In addition,
when
S. pneumoniae 5303 was passaged in
cefuroxime,
it became 4-fold less susceptible to cefaclor,
8-fold less susceptible
to cefpodoxime and loracarbef, 16-fold less
susceptible to cefixime,
and 32-fold less susceptible to
cefuroxime, whereas the susceptibility
of this culture to amoxicillin
and amoxicillin-clavulanate was
unchanged (Table
3).
Time-kill studies with concentrations achievable in MEF.
When
tested at peak concentrations measured in the MEF of children following
the administration of a conventional oral dose (Table 1), cefixime was
inactive against all three strains of Penr
S. pneumoniae tested and cefaclor and
loracarbef only marginally inhibited growth (Fig.
2). Cefprozil had activity similar to
those of cefaclor and loracarbef against S. pneumoniae R1 and S. pneumoniae R2 (MICs, 8 and 16 µg/ml, respectively) but had an antibacterial effect against
S. pneumoniae N1387, which was more susceptible to
cefprozil by agar dilution MIC determinations (MIC, 2 µg/ml). Cefuroxime was marginally more active than cefprozil against
S. pneumoniae R1 and S. pneumoniae R2
(MICs, 4 and 8 µg/ml, respectively) and was effective against
S. pneumoniae N1387 (MIC, 2 µg/ml). In contrast,
the use of amoxicillin and amoxicillin-clavulanate resulted in 2- to 3-log decreases in viable bacterial
numbers for all three strains of Penr S. pneumoniae (MICs, 2 µg/ml) over the 9-h test period.
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FIG. 2.
Bactericidal activities of
amoxicillin-clavulanate ( ), amoxicillin ( ),
cefuroxime ( ), cefprozil ( ), cefaclor ( ), loracarbef
( ), and cefixime ( ) at concentrations achieved in MEF. The growth
in the untreated control culture (×) was used for comparison. (a)
S. pneumoniae N1387. (b) S. pneumoniae
R1. (c) S. pneumoniae R2.
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In vitro pharmacodynamic model.
The MIC data indicated that
amoxicillin, amoxicillin-clavulanate, cefpodoxime, and
cefuroxime are the most potent oral -lactam antibiotics against
S. pneumoniae, including penicillin-resistant strains. These four antibiotics were therefore chosen to be
tested in the in vitro pharmacodynamic model. Twice-daily doses
of amoxicillin (500 mg), amoxicillin-clavulanate (500 plus 125 mg, respectively), cefpodoxime (200 mg), and cefuroxime (250 mg) were simulated against the typically susceptible strain
S. pneumoniae ATCC 6303 (Fig. 3). All were rapidly bactericidal, with
the numbers of viable bacteria being reduced to the limit of detection
(1.67 × 101 CFU/ml) by 6 h in the cultures
treated with cefpodoxime and cefuroxime and by 24 h in the
cultures treated with amoxicillin and
amoxicillin-clavulanate. Some regrowth was seen by 24 h in the cefpodoxime-treated culture, but the viable bacterial count
had fallen below the limit of detection by 26 h and no further
regrowth was seen. No selection of resistance was seen in this study.
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FIG. 3.
Bactericidal activities of simulated concentrations
achieved in the serum of humans following the administration of
oral doses of amoxicillin-clavulanate at 500 plus 125 mg,
respectively, twice daily (), 500 mg of amoxicillin twice daily
(), 200 mg of cefpodoxime twice daily ( ), and 250 mg of
cefuroxime twice daily () against S. pneumoniae ATCC
6303. The growth of an untreated control culture (×) was used for
comparison. Arrows represent the times at which the antibiotic doses
were administered.
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Higher dosages of cefuroxime (500 mg twice daily) and
amoxicillin-clavulanate (875 plus 125 mg, respectively,
twice daily)
were simulated against
S. pneumoniae
1320b, a penicillin-resistant
strain, since higher doses of oral
-lactam antibiotics are recommended
in some countries for
respiratory tract infections. Cefpodoxime
was again tested at 200 mg
twice daily because a higher dose of
this antibiotic is not available.
The concentrations of antibiotic
achieved in the serum of humans
(
18,
19,
34a,
34a,
35)
are presented in Fig.
4. These were simulated in the in vitro
pharmacodynamic model, and microbiological assay results confirmed
that
the antibiotic concentrations obtained in the culture flasks
were
close to those achieved in humans.
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FIG. 4.
Concentrations achieved in the serum of humans following
the administration of oral doses of 875 mg of amoxicillin ()
(34a), 500 mg of amoxicillin () (19), 200 mg
of cefpodoxime () (35), and 500 mg of cefuroxime ()
(18). The MICs of cefpodoxime and cefuroxime
(---) and amoxicillin and
amoxicillin-clavulanate
(....) for S. pneumoniae 1320b are also indicated.
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Simulated concentrations of cefpodoxime achieved in the serum of humans
following the administration of an oral dose of 200
mg were essentially
ineffective, with the growth being similar
to that in the untreated
control culture (Fig.
5). This result
was
not unexpected, because the MIC of cefpodoxime for
S. pneumoniae 1320b is 4 µg/ml, whereas the peak concentration
achievable in
serum following the administration of a 200-mg oral dose
is only
2.1 µg/ml (Fig.
4). The MIC of cefuroxime for this strain is
also
4 µg/ml, but following the administration of an oral dose of 500
mg of cefuroxime, a peak concentration of 8 µg/ml is achieved
in
the serum of humans. Consequently, cefuroxime showed initial
bactericidal activity, but the culture regrew fully between the
doses,
which is consistent with the concentration in serum falling
below 4 µg/ml between 3 and 4 h after dosing (Fig.
4).
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FIG. 5.
Bactericidal activities of simulated concentrations
achieved in the serum of humans following the administration of oral
dosages of amoxicillin-clavulanate at 875 plus 125 mg,
respectively, twice daily, (), 500 mg of amoxicillin twice
daily (), 200 mg of cefpodoxime twice daily (), and 500 mg of
cefuroxime twice daily () against S. pneumoniae
1320b. The growth of an untreated control culture (×) was used for
comparison. Arrows represent the times at which the antibiotic
doses were administered.
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Amoxicillin and amoxicillin-clavulanate showed similar
bactericidal activities up to 6 h, after which the culture treated
with the lower dose of amoxicillin (500 mg) showed some regrowth
by
24 h. The bacterial numbers were reduced to the limit of detection
by 26 h, 2 h after administration of the 24-h dose, but the
amoxicillin-treated
culture fully regrew by 54 h.
Amoxicillin-clavulanate (875 and
125 mg,
respectively) was rapidly bactericidal and reduced the
numbers of
viable bacteria to the limit of detection (1.67 × 10
1
CFU/ml) by 25 h, with no regrowth by the end of the experiment
(54 h) (Fig.
5).
On testing of the organisms isolated at the end of the study for
-lactam susceptibility, cultures of
S. pneumoniae
1320b
treated with simulated serum cefpodoxime and cefuroxime
concentrations
were shown to contain from 48 h onward isolates
that were four-
to eightfold less susceptible to cefpodoxime and
eightfold less
susceptible to cefuroxime than isolates in the
untreated control
culture (Table
4).
These isolates showed cross-resistance with
another
cephalosporin, cefotaxime, but no cross-resistance with
the
penicillins penicillin G and
amoxicillin-clavulanate. The
organisms isolated
following 54 h of exposure to simulated serum
amoxicillin
concentrations and 24 h of exposure to
amoxicillin-clavulanate
showed no decrease in
susceptibility to penicillins or cephalosporins
compared with the
susceptibility of the untreated control culture
(Table
4).
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TABLE 4.
MICs for S. pneumoniae 1320b following
treatment with simulated concentrations of amoxicillin,
amoxicillin-clavulanate, cefuroxime, and cefpodoxime
achieved in the serum of humans
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DISCUSSION |
A number of Peni and Penr strains of
S. pneumoniae were fully susceptible to amoxicillin and
amoxicillin-clavulanate in agar dilution MIC
determinations according to the current breakpoints of the
National Committee for Clinical Laboratory Standards (MICs, 
0.5
µg/ml), whereas some Pens strains were resistant (MICs,
0.5 to 4 µg/ml) to the cephalosporins.
The most potent cephalosporins tested were cefpodoxime and cefuroxime,
but these were also the compounds with which less susceptible isolates
of Pens S. pneumoniae were selected
following repeated subculture. The 4- to 32-fold decreases in
susceptibility to cefpodoxime, cefuroxime, and cefixime seen following
exposure to cefpodoxime and cefuroxime, the lack of cross-resistance
with amoxicillin and amoxicillin-clavulanate, and the fact
that repeated subculture in amoxicillin or
amoxicillin-clavulanate did not lead to the
selection of less susceptible isolates were all consistent with
data reported by Sifaoui et al. (34). Those investigators
isolated one-step mutants from two Pens strains of
S. pneumoniae following growth in the
presence of cefuroxime, cefpodoxime, cefixime, cefotaxime,
and ceftriaxone at frequencies ranging from 1 in 108
to 1 in 106, and the strains were 2- to 16-fold less
susceptible to these agents (34). In the same study,
the frequencies of mutation following exposure to ampicillin,
amoxicillin, amoxicillin-clavulanate, ampicillin-sulbactam,
cefaclor, and loracarbef were up to 10-fold lower, and at most,
only a 2-fold increase in the MIC was observed. Moreover, one class of
cephalosporin-resistant mutants was more susceptible to the penicillins
than the parent cultures (34).
Otitis media is an important community-acquired respiratory tract
infection that is often caused by Penr S. pneumoniae. Recent studies (4, 7, 10) have shown that -lactam resistance has resulted in the clinical and bacteriological failure of oral cephalosporins in the treatment of otitis media, particularly when the MICs for S. pneumoniae exceed 0.5 µg/ml. In our studies, only amoxicillin and
amoxicillin-clavulanate produced a decrease in the numbers
of viable bacteria (99 to 99.9%) for all three strains of
S. pneumoniae tested; this was not surprising since
only these agents achieved peak concentrations in MEF which exceeded
the MICs for all three strains. Cefpodoxime was not tested because data
on its concentration in MEF were not available at the time, although a
recent report (9) has shown that the concentrations of
this compound in MEF (0.2 µg/ml) are at least 10-fold lower than the
MICs for Penr S. pneumoniae. In
addition to the concentrations in MEF, the concentrations of
-lactams in serum have been found to be important in the
bacteriological eradication of the two key pathogens associated with
otitis media, S. pneumoniae and H. influenzae (9), with time above the MIC being the most
significant pharmacokinetic parameter. A time above the MIC in serum of
at least 40% of the dosing interval is required for -lactams to
produce maximal (
80%) bacteriological cure rates for patients with
otitis media and maximal survival rates (90%) in animal models of
infection (8).
The relevance of the time above the MIC in serum was demonstrated in
the in vitro pharmacodynamic model with the four most potent agents,
amoxicillin, amoxicillin-clavulanate, cefpodoxime, and
cefuroxime, against Pens and Penr strains of
S. pneumoniae. The model simulates the
concentrations of antibiotic achieved in the serum of humans,
considered by many investigators to be predictive of outcome in
the treatment of respiratory tract infections caused by pathogens, such
as S. pneumoniae and H. influenzae,
which are not intracellular (5, 12, 30). The rapid
bactericidal activity and lack of selection of resistance seen against
S. pneumoniae ATCC 6303 were most likely because of the
long period of time above the MICs of the antibiotics tested (100% of
the dosing interval). The lack of selection of resistance seen with
this strain in the pharmacodynamic model was consistent with the fact
that the development of -lactam resistance by Pens
S. pneumoniae in the clinic is considered to have
originally arisen via acquisition of genetic material from other
streptococcal species rather than by mutation within S. pneumoniae (11).
It has been suggested, however, that once the mosaic genes encoding
less susceptible PBPs have been acquired by S. pneumoniae, they may mutate further to give even higher levels of
-lactam resistance (2, 6). This was examined in the in
vitro model, in which penicillin-resistant strain S. pneumoniae 1320b was also exposed to amoxicillin,
amoxicillin-clavulanate, cefpodoxime, and cefuroxime at
concentrations achieved in the serum of humans. Although the
concentrations of amoxicillin and cefuroxime obtained following the
administration of an oral dose of 500 mg are similar, the lower MIC of
amoxicillin meant that the time above the MIC was longer for this
antibiotic (7 h in 24 h for amoxicillin, 5 h in 24 h for
cefuroxime). Even so, regrowth of S. pneumoniae was
seen between doses for both amoxicillin and cefuroxime but not for mg
amoxicillin-clavulanate at 875 and 125 mg, respectively. This was probably due to the increased time above the MIC for the
amoxicillin component, which was 10 of 24 h (42%) for the dosage
of 875 mg twice daily.
As seen in the passage experiments, isolates with reduced
cephalosporin susceptibility emerged following exposure of
S. pneumoniae 1320b to simulated serum cefpodoxime
and cefuroxime concentrations. This was unexpected in the case of
cefpodoxime, because there appeared to be no selection pressure for a
higher level of resistance, since the MIC for the parent culture of
S. pneumoniae 1320b (4 µg of cefpodoxime/ml) already
exceeded the maximum concentration in serum (2.1 µg of
cefpodoxime/ml). The finding was consistent, however, with data
reported by Negri et al. (29) in 1994. They showed
that when populations of Peni and Penr
S. pneumoniae were mixed, the Penr strain
was selected as prevalent by all the -lactams tested, even at
concentrations below the MIC for the Peni strain. In our
study, the isolates selected from cultures treated with simulated serum
cefpodoxime and cefuroxime concentrations were four- to eightfold less
susceptible to cefpodoxime, cefuroxime, and cefotaxime but were not
cross-resistant with the penicillins tested. This is consistent with
mutations in PBPs 1a and 2x, which have been shown to be necessary for
cephalosporin resistance in S. pneumoniae, and was
exemplified by a characterized variant, S. pneumoniae
R61a2x (6). Mutation in PBP 2b is also required to confer
resistance to penicillins. The decreased susceptibility to cefotaxime
observed in the present study (MICs of up to 8 µg/ml) could have
severe consequences in the treatment of more serious infections caused
by S. pneumoniae, such as meningitis, for which parenteral cephalosporins are often recommended when Penr
pneumococci are suspected.
In conclusion, cross-resistance between -lactam antibiotics cannot
be assumed for S. pneumoniae, and in the clinical
situation, it is important to determine the susceptibility of the
strain to the specific agent being considered for use in treatment.
Because this is not likely to be known when empirical therapy of
community-acquired infections is required, epidemiological data and
local resistance patterns must be relied upon and the susceptibilities
of other important respiratory pathogens, such as H. influenzae and M. catarrhalis, which may produce
-lactamases, should also be considered. It is important to choose an
agent that is unlikely to lead to the selection of a higher level of
resistance and to administer it at a dose high enough to ensure optimal
coverage against penicillin-resistant pneumococci. In these studies,
amoxicillin and amoxicillin-clavulanate were the most effective oral -lactams against penicillin-susceptible and penicillin-resistant pneumococci and, unlike some of the
cephalosporins, did not lead to the selection of resistance.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: c/o Tony White,
Anti-infectives SPD, SmithKline Beecham Pharmaceuticals, New Frontiers Science Park (South), Third Avenue, Harlow, Essex CM19 5AW, United Kingdom. Phone: 01279 644373. Fax: 01279 646039.
| |
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Top
Abstract
Introduction
