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Antimicrobial Agents and Chemotherapy, May 2000, p. 1377-1380, Vol. 44, No. 5
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Comparative Killing Rates of Fluoroquinolones
and Cell Wall-Active Agents
Joan C.
Fung-Tomc,*
Elizabeth
Gradelski,
Lourdes
Valera,
Benjamin
Kolek, and
Daniel P.
Bonner
Department of Microbiology, Bristol-Myers
Squibb Company, Wallingford, Connecticut 06492
Received 8 October 1999/Returned for modification 24 November
1999/Accepted 11 February 2000
 |
ABSTRACT |
Killing rates of fluoroquinolones, 
-lactams, and vancomycin were
compared against Enterobacteriaceae, Staphylococcus
aureus, pneumococci, streptococci, and Enterococcus
faecalis. The times required for fluoroquinolones to decrease
viability by 3 log10 were 1.5 h for
Enterobacteriaceae, 4 to 6 h for staphylococci, and
6 h for streptococci and enterococci. Thus, the rate of killing by
fluoroquinolones is organism group dependent; overall, they killed more
rapidly than -lactams and vancomycin.
| |
TEXT |
The ability of an antibiotic to kill
bacteria may be important in some clinical settings, such as the
management of bacterial endocarditis or the treatment of bacteremia in
granulocytopenic patients. Members of the primary antimicrobial classes
showing bactericidal properties that are used therapeutically as single agents are the quinolones and the cell wall-active agents, specifically -lactams and the glycopeptide vancomycin.
Quinolones exhibit concentration-dependent killing kinetics, with
maximum killing rates achieved at the optimum bactericidal concentration (12), which for more recently developed
fluoroquinolones occurs close to eight times their MICs
(E. Gradelski, B. Kolek, D. Bonner, and J. Fung-Tomc, Abstr.
38th Intersci. Conf. Antimicrob. Agents Chemother., abstr. E-182,
1998). On the other hand, -lactam antibiotics and vancomycin
exhibit time-dependent kinetics (8), with little difference
in killing rates at concentrations above their MICs.
In this study, we compared the killing rates of three
fluoroquinolones (trovafloxacin, levofloxacin, and gatifloxacin)
to those of cell wall-active agents commonly used in the
treatment of infections involving specific bacterial species.
Bacterial strains used in this study were clinical isolates: four
members of the family Enterobacteriaceae, four
Staphylococcus aureus isolates (two methicillin susceptible
and two methicillin resistant), four streptococci (two alpha-hemolytic
and two beta-hemolytic streptococci), nine Streptococcus
pneumoniae isolates (with various penicillin susceptibilities),
and five Enterococcus faecalis isolates. The strains were
chosen because their quinolone MICs were close to the respective
quinolone-bacterial species modal MICs.
Gatifloxacin was from Kyorin Pharmaceutical Co. (Tochigi, Japan),
methicillin was from Bristol-Myers Squibb Co. (Wallingford, Conn.),
trovafloxacin was from Pfizer Inc. (New York, N.Y.), and levofloxacin
was from the R. W. Johnson Pharmaceutical Research Institute
(Raritan, N.J.). Ceftriaxone was supplied by Roche Pharmaceuticals (Nutley, N.J.), cefotaxime was supplied by Hoechst Marion Roussel (Kansas City, Mo.), and vancomycin was supplied by Eli Lilly & Co.
(Indianapolis, Ind.). Penicillin G, amoxicillin, and ampicillin were
purchased from Sigma Chemical Co. (St. Louis, Mo.).
MICs were determined by the agar dilution method with Mueller-Hinton
agar (supplemented with 5% sheep blood for pneumococci) according to
standardized methods (13). The MIC is defined as the lowest
concentration of antibiotic that inhibited all visible growth.
Time-kill kinetics were conducted at drug concentrations equal to 10 times the MIC of that drug for a bacterial strain. The optimal
bactericidal concentrations of quinolones are close to eight times
their MICs. The maximum killing rates of seven -lactams and
vancomycin for seven strains used in this study were determined (data
not shown); maximum killing occurred at four to eight times the MIC of
a drug, and this rate was maintained up to 16 times the MIC. The tests
were performed in Mueller-Hinton broth (staphylococci, Escherichia coli, and E. faecalis) which, for
S. pneumoniae and viridans streptococcal strains, was
supplemented with 7% lysed horse blood. The beta-hemolytic
streptococcal time-kill studies were done in brain heart infusion
broth, a medium yielding better growth of these organisms.
Twenty-milliliter cultures, grown in 50-ml glass flasks, were incubated
at 35°C with shaking. Cells were grown to logarithmic phase with
1 h of preincubation in fresh broth prior to the addition of drug.
The starting bacterial density was approximately 5 × 105 to 1 × 106 CFU/ml. Viable counts were
determined at 0, 2, 4, 6, and (except for pneumococci) 24 h after
drug addition by plating known dilutions of the samples onto
Mueller-Hinton agar (or for streptococci, Trypticase agar plus 5%
sheep blood). The cell count plates were incubated for up to 48 h
before any were considered to have no growth. With a number of strains,
the cell counts were simultaneously determined with the agar plates
described above supplemented with 1% magnesium chloride (M. Wooton,
K. E. Bowker, H. A. Holt, and A. P. MacGowan,
Abstr. 38th Intersci. Conf. Antimicrob. Agents Chemother.,
abstr. A-32, 1998), a condition known to diminish the effect of any
quinolone carryover. In all cases, the bacterial cell counts for these
strains were the same on both plating media. Also, plating of the
culture immediately after the addition of quinolone resulted in the
same cell count as that of the culture plated immediately prior to the
addition of drug.
Results and discussion.
MICs and the rates of killing
nonstreptococcal species are listed in Table
1. Fluoroquinolone MICs for the
Enterobacteriaceae and staphylococci were 
0.13 and 0.25
µg/ml, respectively.
Since a 3 log
10 drop in viability is considered an index of
bactericidal activity (
14), the time required to achieve
this
level of killing was determined from time-kill curves. There were
minor differences among the fluoroquinolones in their rates of
killing.
They tended to kill
Enterobacteriaceae (i.e.,
E. coli,
Klebsiella pneumoniae, and
Enterobacter
cloacae) very rapidly,
achieving a 3 log
10 decrease by
1.5 h (Table
1). In comparison,
the fluoroquinolones killed
staphylococci more slowly, with 3
log
10 killing by 4 to
6 h of exposure. Three of the four staphylococcal
strains, exposed
to gatifloxacin, showed a 3 log
10 decrease in
viability by
2 h. The differential killing rates of the
Enterobacteriaceae and
S. aureus by
fluoroquinolones are displayed in Fig.
1.
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FIG. 1.
Time-kill curves for nonstreptococcal species: E. coli A20697 (A), K. pneumoniae A27464 (B), and
MSSA A9606 (C).
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|
Others have reported the more rapid killing of gram-negative bacteria
by quinolones. Pefloxacin was more rapidly bactericidal
against
gram-negative strains in the first hour of contact, but
the rates of
killing of gram-negative and gram-positive strains
were similar by
4 h of incubation (
3). Likewise, PD 131628
was more
bactericidal against
E. coli than against staphylococci
(
9).
The cell wall-active agents were less rapidly bactericidal than the
fluoroquinolones. To attain a 3 log
10 drop in viability
against
Enterobacteriaceae strains, 6 to >24 h of exposure
to
cefotaxime was needed (Table
1). Regrowth was noted in three
of the
cefotaxime-treated
Enterobacteriaceae cultures by 24 h
of incubation, but heavy regrowth with
E. cloacae A20650 was
observed.
Although the reason for the regrowth was not explored, with
E. cloacae A20650, cefotaxime-resistant variants (i.e.,
cefotaxime
MICs of >16 µg/ml) accounted for the heavy regrowth. Some
extended-spectrum
cephalosporins can readily select for resistant
variants of
E. cloacae (
6). Methicillin and
cefotaxime killed methicillin-susceptible
S. aureus (MSSA)
strains A9606 and A15090 more slowly (4 to 6
h for 3 log
10 killing), as did vancomycin against the
methicillin-resistant
S. aureus (MRSA) strains (>6 to
24 h for 3 log
10 killing). Thus,
fluoroquinolones were
more rapidly bactericidal than cefotaxime
against MSSA and
Enterobacteriaceae, more rapid than methicillin
against
MSSA, and more rapid than vancomycin against MRSA
strains.
Compared to nonstreptococcal species, fluoroquinolones were
less rapidly lethal against the 13 streptococcal and 5 enterococcal
strains. Both fluoroquinolones and cell wall-active agents
required
>6 to >24 h to reduce viability by 3 log
10. On
the whole, all
of the drugs tested killed streptococci and
enterococci more slowly
than the nonstreptococcal strains (Fig.
2).
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|
FIG. 2.
Time-kill curves for streptococci and related species:
penicillin-susceptible S. pneumoniae A9585 (A), S. mitis A28590 (B), and S. pyogenes A20789 (C).
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The data in this study on quinolone killing of streptococci are
more extensive than, but confirmatory of, previous reports
(Table
2). Moxifloxacin was reported to be most
rapidly bactericidal
for
S. aureus and
E. coli and least bactericidal for
Streptococcus pyogenes
(
1,
15). Trovafloxacin was bactericidal against
MRSA strains
but killed
Streptococcus mitis and
Streptococcus sanguis much more slowly (
5). A low rate of
enterococcal killing
had been reported for ciprofloxacin (
10,
12), DU-6859a (i.e.,
sitafloxacin) (
12), trovafloxacin
(
11,
15), DR-3355 (
10),
and PD 131628 (
9).
A comparison of killing rates by ampicillin and moxifloxacin of
E. faecalis (F. Maggiolo, R. Capra, P. Bottura, M. Moroni,
G. Pravettoni, and F. Suter, Abstr. 38th Intersci. Conf. Antimicrob.
Agents Chemother., abstr. E-201, 1998), by ceftriaxone and sparfloxacin
of viridans streptococci (
4), and by penicillin and
temafloxacin
of
Streptococcus adjacens (
2)
suggests that the
-lactams may
kill streptococci and
enterococci even more slowly than do quinolones.
Recently, Hoellman et
al. (
7) reported the killing rates of
12 pneumococcal
strains by fluoroquinolones and
-lactams; at
eight times the MICs of
the drugs, five to eight strains showed
3 log
10 killing at
6 h by ciprofloxacin, levofloxacin, gatifloxacin,
sparfloxacin,
and trovafloxacin; five to six strains showed killing
at 6 h
by amoxicillin, cefuroxime, and ceftriaxone. For all 12
pneumococcal
strains to show a 3 log
10 decrease in cell counts,
the
quinolones required 12 h of contact, while the
-lactams needed
24 h for an equal effect (
7).
In summary, these results indicate that the rate of killing by
fluoroquinolones is bacterial group dependent. Fluoroquinolones
kill
gram-negative bacteria more rapidly than staphylococci. Quinolones
killed nonstreptococcal strains more rapidly than did
-lactams
and
vancomycin. Although fluoroquinolones kill streptococci and
enterococci
more slowly, these rates may be similar to, and possibly
higher
than, those achieved by the
-lactams and vancomycin. Overall,
against a spectrum of gram-positive and gram-negative bacteria,
the quinolones exhibit the more favorable bactericidal
profile.
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FOOTNOTES |
*
Corresponding author. Mailing address: Bristol-Myers
Squibb Company, Department of Microbiology-104, 5 Research Pkwy.,
Wallingford, CT 06492. Phone: (203) 677-6370. Fax: (203) 677-6771. E-mail: fungtomj{at}bms.com.
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Antimicrobial Agents and Chemotherapy, May 2000, p. 1377-1380, Vol. 44, No. 5
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
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