ANTIMICROBIAL AGzNT AND CHEMOTHERAPY, Feb. 1977, Copyright © 1977 American Society for Microbiology
p.
258-261
Vol. 11, No. 2 Printed in U.S.A.
In Vitro Activity of Ticarcillin Against Anaerobic Bacteria Compared with That of Carbenicillin and Penicillin IRA ROY, VINH BACH, AND HARAGOPAL THADEPALLI* Division of Infectious Diseases* and Department ofInternal Medicine, Martin Luther King, Jr. General Hospital; Charles R. Drew Postgraduate Medical School; and University of Southern California School of Medicine, Los Angeles, California 90059 Received for publication 8 September 1976
A total of 334 clinical anaerobic isolates were tested in an anaerobic glove box by the agar dilution technique for susceptibility to clinically achievable levels of ticarcillin, carbenicillin, and penicillin. Thirty-two micrograms or less of penicillin per milliliter inhibited 91% of all strains, whereas 100 ug of carbenicillin and ticarcillin per ml inhibited 95 and 98%, respectively. A total of 82% (85 strains) of Bacteroides were inhibited by penicillin, and 93 and 96% were inhibited by carbenicillin and ticarcillin, respectively. Thirteen (24%) of 55 strains of Bacteroides fragilis tested were resistant to 32 ,ug of penicillin per ml, and 6 (11%) and 3 (5%) were resistant to 100 ug of carbenicillin and ticarcillin per ml, respectively. Within the therapeutic range, ticarcillin was the most effective of the three penicillins tested against B. fragilis subsp. fragilis.
Ticarcillin, a new semisynthetic penicillin, resembles carbenicillin in structure and in in vitro antimicrobial activity against aerobic bacteria including Pseudomonas species. Its effect against anaerobic bacteria is not established. We examined the efficacy of ticarcillin against 334 clinical anaerobic isolates (234 ofthem were derived from the female genital tract) in comparison with the effect of penicillin and carbenicillin in vitro. (This work was presented in part at the 76th Annual Meeting of the American Society for Microbiology, Atlantic City, N.J., 2-7 May 1976.) MATERIALS AND METHODS Identification of all isolates was done as described in the anaerobic laboratory manual of the Virginia Polytechnic Institute (1). All 334 were human isolates identified in our laboratory between August 1975 and July 1976. Two-thirds of these (i.e., 234) were isolated from healthy cervices or from female genital tract infections, and the remaining were from infections of the lung or abdomen. The inocula were overnight brain heart infusion broth cultures diluted to McFarland standard I delivered with a Steers replicator to deposit 105 colony-forming units. Solutions containing concentrations of drug were prepared by dissolving the standard powders of penicillin G (Eli Lilly & Co., Indianapolis, Ind.), sodium carbenicillin (Roerig), and disodium ticarcillin (Beecham Products Inc., Clifton, N.J.) in distilled water. The antibiotic solutions were freshly prepared and added to brain heart infusion agar (Clinical Standard, Torrance, Calif.) and supplemented
with defibrinated sheep blood (5%), 10 ug of vitamin K per ml, and hemin (Scott Laboratories, Fiskeville, R. I.). The inocula were planted with a Steers replicator (6), and the plates were incubated in an anaerobic glove box filled with a gas mixture of 85% nitrogen, 5% carbon dioxide, and 10% hydrogen and incubated at 37°C for 48 h. The minimal inhibitory concentration (MIC) was read as the least amount of antibiotic that permitted no growth. A set of aerobic and anaerobic plates without antibiotics was included as a control with each set of tests.
RESULTS Tables 1, 2, and 3 depict the MIC of penicillin, carbenicillin, and ticarcillin for inhibition of 334 anaerobic isolates. All three penicillins inhibited more than 90% of the anaerobes tested. As expected, 25% of the B. fragilis subsp. fragilis was resistant to 32 ,ug of penicillin per ml; correspondingly, 18 and 7% were also resistant to carbenicillin and ticarcillin, respectively. Three of 28 (11%) strains resistant to both penicillin and carbenicillin, but susceptible to ticarcillin, accounted for this difference. Two strains (7%) of B. fragilis subsp. fragilis were resistant to all three penicillins. B. fragilis, other than subsp. fragilis, were inhibited by both carbenicillin and ticarcillin alike, but 22% of them were resistant to penicillin. Strains of the Bacteroides group, except B. fragilis, were highly susceptible to carbenicillin (100%) and ticarcillin (97%). In general, 18% of all strains ofBacteroides were resistant to peni258
TICARCILLIN AGAINST ANAEROBIC BACTERIA
VOL. 1 l, 1977
259
TABLE 1. Penicillin concentration and cumulative percentage of susceptible strains Organisms
No. of strains
Gram-negative rods Bacteroides (B) fragilis subsp. fragilis B. fragilis other than B. fragilis subsp. fra-
%
Susceptible at MIC (g.g/ml) of:
2
4
8
16
32
64
28 27
32 26
36 41
52
54 59
75 78
96 100
30 (85) 10 1 44
77 46 60 100 52
80 53
83 54
93 69 70
82
97 97 80
55
86
95
100
Nonsporeforming gram-positive rods Eubacterium Lactobacilli Propionibacteria Bifidobacteria Actinomyces
36 16 8 3 7
67 88 75 100 100
72
75 94
89
92
Cocci Peptococci Peptostreptococci Veillonella Megasphera elsdenii
75 43 4 2
77 49
85 86 50 100
88 88
93 93
334
(60)
(70)
(77)
(84)
gilis
Bacteroides except B. fragilis Bacteroides (all strains) Fusobacterium Selanomonas Clostridia (all)
Total (avg)
95 100
(91)
95 94 75
93 95
(96)
TABLE 2. Carbenicillin concentration and cumulative percentage of susceptible strains Organisms
No. of strains
5
of: %10Susceptible 25 at MIC50(ug/ml)100
200
Gram-negative rods Bacteroides (B) fragilis subsp. fragilis B. fragilis other than B. fragilis subsp. fra-
28 27
36 56
54
30 10 1 44
93 62 90 100 52
36 16 8 3 7
83 87 75 100 71
75 43 4
100
2
89 86 25 100
334
(75)
(77)
gilis
Bacteroides except B. fragilis Bacteroides (all above strains) Fusobacteria Selanomonas Clostridia (all)
Nonsporeforming gram-positive rods Eubacteria Lactobacilli Propionibacteria Bifidobacteria Actinomyces
(85)
67
71 89
82 96
100 100
100 71
79
87
93
100 90
55
91
95
97
97
86
89
97 100
100
68
75 100
Cocci
Peptococci Peptostreptococci
Veillonella Megasphera elsdenii
Total (avg)
97
94
100 (85)
(91)
(96)
(98)
260
ROY, BACH, AND THADEPALTLI
Ai-mmicRoB. AGzNTs CHEMOTHZR. TABLE 3. Ticarcillin concentration and cumulative percentage of susceptible strains No. of
Org sstrains
5
% Susceptible at MIC (,ug/ml) of: 10 25 50 100
Gram-negative rods Bacteroides (B) fragilis subsp. fragilis B. fragilis other than B. fragilis subsp. fragilis Bacteroides except B. fragilis Bacteroides (all above strains) Fusobacteria Selanomonas Clostridia (all)
28 27
39 67
46
50 70
71 93
30 (85) 10 1 44
93 67 80 100 59
97 71
73
100 88
66
86
93
Nonsporeforming gram-positive rods Eubacteria Lactobacilli Propionibacteria Bifidobacteria Actinomyces
36 16 8 3 7
94 88 75 100 71
97
75 43 4 2
60 88 100 100
98 93
98
334
(77)
(84)
(88)
Cocci Peptococci Peptostreptococci
Veillonella Megasphera elsdenii Total (avg)
200
93 96
100 100
96 100
100
100
100 100
75 100 99
(94)
(98)
(99)
cillin, and 7 and 4% were resistant to carbeni- find a place in the treatment of postsurgical infections of the female genital tract. cillin and ticarcillin, respectively. Although all three penicillins exhibited more Forty-four strains of ten different species of clostridia were susceptible to all three penicil- than 90% efficacy against 334 anaerobic lins. Twenty-five (57%) of these strains were of strains, there were important differences in species C. ramosum. Actinomyces (seven their effectiveness against the Bacteroides strains), including four strains of A. israellii group of organisms. Both carbenicillin and tiand 3 strains of A. naeslundii, were suscepti- carcillin inhibited 93 to 96% of all strains of ble to all three penicillins. Most gram-positive Bacteroides, including B. fragilis other than B. cocci with occasional exceptions were also sus- fragilis subsp. fragilis. Thirty-two micrograms of penicillin per ml was ineffective against 18% ceptible to all three penicillins. of these strains. B. fragilis subsp. fragilis, howwere the most resistant of all Bacteroides. ever, DISCUSSION It is difficult to treat a deep-seated infection Ticarcillin, like carbenicillin (also a semisyn- with penicillin G when the MICs of the infectthetic penicillin), was found to be effective ing organisms are higher than 25 gg/ml (7). against aerobic bacteria such as Pseudomonas Hence, the cutoff point chosen for penicillin was aeruginosa both in vitro and in vivo (2-5, 8). It 32 ,ug/ml. Even at this concentration, nearly was reported that ticarcillin might even have a 25% of the strains of B. fragilis subsp. fragilis greater spectrum of activity and might achieve were resistant to penicillin. Both carbenicillin relatively higher drug concentrations in serum and ticarcillin inhibited 82 and 93% of these than carbenicillin (5, 8). Its effect on anaerobes strains, respectively. The relative resistance of some ofthe isolates in vitro is not well known. The in vivo efficacy of this drug is now being tested. We chose 100 to carbenicillin while susceptible to ticarcillin gg/ml as a cutoff point for ticarcillin because may be real or may be a spurious reflection of the reported concentrations of this drug in the increased usage of carbenicillin in our hosserum 0.5 to 1.0 h after a 5-g intravenous injec- pital in recent years. This remains to be verition were found to be from 200 ug (4) to as high fied. as 740 + 56 ,mg/ml (5). Nearly two-thirds of the LITERATURE CITED strains reported here were from the female gen1. Holdeman, L. V., and W. E. C. Moore (ed.). 1975. Anaerobe laboratory manual. Virginia Polytechnic Inital tract, which suggests that ticarcillin may
VOL. 11, 1977
TICARCILLIN AGAINST ANAEROBIC BACTERIA
stitute Anaerobe Laboratory, Virginia Polytechnic Institute and State University, Blacksburg. 2. Neu, H. C., and G. J. Garvey. 1975. Comparative in vitro activity and clinicalipharmacology of ticarcillin and carbenicillin. Antimicrob. Agents Chemother. 8:457-462. 3. Pines, A., G. Khaja, H. Raafat, and K. S. Shreedharan. 1974. Preliminary clinical experience with Ticarcillin (BRL2288) in 101 patients tteated for severe respiratory infections. Chemotherapy 20:3944. 4. Rodriguez, V., G. P. Bodey, N. Horikoshi, J. Inagaki, and K. B. McCredie. 1973. Ticarcillin therapy of infections. Antimicrob. Agents Chemother. 4:427431.
261
5. Simon, C., M. Leuth, and V. Malerczyk. 1974. Ticarcillin. Deutsch. Med. Woschenschr. 99:2460-2464. 6. Steers, E., E. L. Foltz, and B. S. Graves. 1959. An inocula replicating apparatus for routine testing of bacterial susceptibiliiy to antibiotics. Antibiot. Chemother. 9:307-311. 7. Sutter, V. L., and S. M. Finegold. 1975. Susceptibility of anaerobic bacteria to carbenicillin, cefoxitin and related drugs. J. Infect. Dis. 131:417422. 8. Wise, R., and D. S. Reeves. 1974. Clinical and laboratory investigations on Ticarcillin and anti-pseudomonal antibiotic. Chemotherapy 20:45-51.
p.
258-261
Vol. 11, No. 2 Printed in U.S.A.
In Vitro Activity of Ticarcillin Against Anaerobic Bacteria Compared with That of Carbenicillin and Penicillin IRA ROY, VINH BACH, AND HARAGOPAL THADEPALLI* Division of Infectious Diseases* and Department ofInternal Medicine, Martin Luther King, Jr. General Hospital; Charles R. Drew Postgraduate Medical School; and University of Southern California School of Medicine, Los Angeles, California 90059 Received for publication 8 September 1976
A total of 334 clinical anaerobic isolates were tested in an anaerobic glove box by the agar dilution technique for susceptibility to clinically achievable levels of ticarcillin, carbenicillin, and penicillin. Thirty-two micrograms or less of penicillin per milliliter inhibited 91% of all strains, whereas 100 ug of carbenicillin and ticarcillin per ml inhibited 95 and 98%, respectively. A total of 82% (85 strains) of Bacteroides were inhibited by penicillin, and 93 and 96% were inhibited by carbenicillin and ticarcillin, respectively. Thirteen (24%) of 55 strains of Bacteroides fragilis tested were resistant to 32 ,ug of penicillin per ml, and 6 (11%) and 3 (5%) were resistant to 100 ug of carbenicillin and ticarcillin per ml, respectively. Within the therapeutic range, ticarcillin was the most effective of the three penicillins tested against B. fragilis subsp. fragilis.
Ticarcillin, a new semisynthetic penicillin, resembles carbenicillin in structure and in in vitro antimicrobial activity against aerobic bacteria including Pseudomonas species. Its effect against anaerobic bacteria is not established. We examined the efficacy of ticarcillin against 334 clinical anaerobic isolates (234 ofthem were derived from the female genital tract) in comparison with the effect of penicillin and carbenicillin in vitro. (This work was presented in part at the 76th Annual Meeting of the American Society for Microbiology, Atlantic City, N.J., 2-7 May 1976.) MATERIALS AND METHODS Identification of all isolates was done as described in the anaerobic laboratory manual of the Virginia Polytechnic Institute (1). All 334 were human isolates identified in our laboratory between August 1975 and July 1976. Two-thirds of these (i.e., 234) were isolated from healthy cervices or from female genital tract infections, and the remaining were from infections of the lung or abdomen. The inocula were overnight brain heart infusion broth cultures diluted to McFarland standard I delivered with a Steers replicator to deposit 105 colony-forming units. Solutions containing concentrations of drug were prepared by dissolving the standard powders of penicillin G (Eli Lilly & Co., Indianapolis, Ind.), sodium carbenicillin (Roerig), and disodium ticarcillin (Beecham Products Inc., Clifton, N.J.) in distilled water. The antibiotic solutions were freshly prepared and added to brain heart infusion agar (Clinical Standard, Torrance, Calif.) and supplemented
with defibrinated sheep blood (5%), 10 ug of vitamin K per ml, and hemin (Scott Laboratories, Fiskeville, R. I.). The inocula were planted with a Steers replicator (6), and the plates were incubated in an anaerobic glove box filled with a gas mixture of 85% nitrogen, 5% carbon dioxide, and 10% hydrogen and incubated at 37°C for 48 h. The minimal inhibitory concentration (MIC) was read as the least amount of antibiotic that permitted no growth. A set of aerobic and anaerobic plates without antibiotics was included as a control with each set of tests.
RESULTS Tables 1, 2, and 3 depict the MIC of penicillin, carbenicillin, and ticarcillin for inhibition of 334 anaerobic isolates. All three penicillins inhibited more than 90% of the anaerobes tested. As expected, 25% of the B. fragilis subsp. fragilis was resistant to 32 ,ug of penicillin per ml; correspondingly, 18 and 7% were also resistant to carbenicillin and ticarcillin, respectively. Three of 28 (11%) strains resistant to both penicillin and carbenicillin, but susceptible to ticarcillin, accounted for this difference. Two strains (7%) of B. fragilis subsp. fragilis were resistant to all three penicillins. B. fragilis, other than subsp. fragilis, were inhibited by both carbenicillin and ticarcillin alike, but 22% of them were resistant to penicillin. Strains of the Bacteroides group, except B. fragilis, were highly susceptible to carbenicillin (100%) and ticarcillin (97%). In general, 18% of all strains ofBacteroides were resistant to peni258
TICARCILLIN AGAINST ANAEROBIC BACTERIA
VOL. 1 l, 1977
259
TABLE 1. Penicillin concentration and cumulative percentage of susceptible strains Organisms
No. of strains
Gram-negative rods Bacteroides (B) fragilis subsp. fragilis B. fragilis other than B. fragilis subsp. fra-
%
Susceptible at MIC (g.g/ml) of:
2
4
8
16
32
64
28 27
32 26
36 41
52
54 59
75 78
96 100
30 (85) 10 1 44
77 46 60 100 52
80 53
83 54
93 69 70
82
97 97 80
55
86
95
100
Nonsporeforming gram-positive rods Eubacterium Lactobacilli Propionibacteria Bifidobacteria Actinomyces
36 16 8 3 7
67 88 75 100 100
72
75 94
89
92
Cocci Peptococci Peptostreptococci Veillonella Megasphera elsdenii
75 43 4 2
77 49
85 86 50 100
88 88
93 93
334
(60)
(70)
(77)
(84)
gilis
Bacteroides except B. fragilis Bacteroides (all strains) Fusobacterium Selanomonas Clostridia (all)
Total (avg)
95 100
(91)
95 94 75
93 95
(96)
TABLE 2. Carbenicillin concentration and cumulative percentage of susceptible strains Organisms
No. of strains
5
of: %10Susceptible 25 at MIC50(ug/ml)100
200
Gram-negative rods Bacteroides (B) fragilis subsp. fragilis B. fragilis other than B. fragilis subsp. fra-
28 27
36 56
54
30 10 1 44
93 62 90 100 52
36 16 8 3 7
83 87 75 100 71
75 43 4
100
2
89 86 25 100
334
(75)
(77)
gilis
Bacteroides except B. fragilis Bacteroides (all above strains) Fusobacteria Selanomonas Clostridia (all)
Nonsporeforming gram-positive rods Eubacteria Lactobacilli Propionibacteria Bifidobacteria Actinomyces
(85)
67
71 89
82 96
100 100
100 71
79
87
93
100 90
55
91
95
97
97
86
89
97 100
100
68
75 100
Cocci
Peptococci Peptostreptococci
Veillonella Megasphera elsdenii
Total (avg)
97
94
100 (85)
(91)
(96)
(98)
260
ROY, BACH, AND THADEPALTLI
Ai-mmicRoB. AGzNTs CHEMOTHZR. TABLE 3. Ticarcillin concentration and cumulative percentage of susceptible strains No. of
Org sstrains
5
% Susceptible at MIC (,ug/ml) of: 10 25 50 100
Gram-negative rods Bacteroides (B) fragilis subsp. fragilis B. fragilis other than B. fragilis subsp. fragilis Bacteroides except B. fragilis Bacteroides (all above strains) Fusobacteria Selanomonas Clostridia (all)
28 27
39 67
46
50 70
71 93
30 (85) 10 1 44
93 67 80 100 59
97 71
73
100 88
66
86
93
Nonsporeforming gram-positive rods Eubacteria Lactobacilli Propionibacteria Bifidobacteria Actinomyces
36 16 8 3 7
94 88 75 100 71
97
75 43 4 2
60 88 100 100
98 93
98
334
(77)
(84)
(88)
Cocci Peptococci Peptostreptococci
Veillonella Megasphera elsdenii Total (avg)
200
93 96
100 100
96 100
100
100
100 100
75 100 99
(94)
(98)
(99)
cillin, and 7 and 4% were resistant to carbeni- find a place in the treatment of postsurgical infections of the female genital tract. cillin and ticarcillin, respectively. Although all three penicillins exhibited more Forty-four strains of ten different species of clostridia were susceptible to all three penicil- than 90% efficacy against 334 anaerobic lins. Twenty-five (57%) of these strains were of strains, there were important differences in species C. ramosum. Actinomyces (seven their effectiveness against the Bacteroides strains), including four strains of A. israellii group of organisms. Both carbenicillin and tiand 3 strains of A. naeslundii, were suscepti- carcillin inhibited 93 to 96% of all strains of ble to all three penicillins. Most gram-positive Bacteroides, including B. fragilis other than B. cocci with occasional exceptions were also sus- fragilis subsp. fragilis. Thirty-two micrograms of penicillin per ml was ineffective against 18% ceptible to all three penicillins. of these strains. B. fragilis subsp. fragilis, howwere the most resistant of all Bacteroides. ever, DISCUSSION It is difficult to treat a deep-seated infection Ticarcillin, like carbenicillin (also a semisyn- with penicillin G when the MICs of the infectthetic penicillin), was found to be effective ing organisms are higher than 25 gg/ml (7). against aerobic bacteria such as Pseudomonas Hence, the cutoff point chosen for penicillin was aeruginosa both in vitro and in vivo (2-5, 8). It 32 ,ug/ml. Even at this concentration, nearly was reported that ticarcillin might even have a 25% of the strains of B. fragilis subsp. fragilis greater spectrum of activity and might achieve were resistant to penicillin. Both carbenicillin relatively higher drug concentrations in serum and ticarcillin inhibited 82 and 93% of these than carbenicillin (5, 8). Its effect on anaerobes strains, respectively. The relative resistance of some ofthe isolates in vitro is not well known. The in vivo efficacy of this drug is now being tested. We chose 100 to carbenicillin while susceptible to ticarcillin gg/ml as a cutoff point for ticarcillin because may be real or may be a spurious reflection of the reported concentrations of this drug in the increased usage of carbenicillin in our hosserum 0.5 to 1.0 h after a 5-g intravenous injec- pital in recent years. This remains to be verition were found to be from 200 ug (4) to as high fied. as 740 + 56 ,mg/ml (5). Nearly two-thirds of the LITERATURE CITED strains reported here were from the female gen1. Holdeman, L. V., and W. E. C. Moore (ed.). 1975. Anaerobe laboratory manual. Virginia Polytechnic Inital tract, which suggests that ticarcillin may
VOL. 11, 1977
TICARCILLIN AGAINST ANAEROBIC BACTERIA
stitute Anaerobe Laboratory, Virginia Polytechnic Institute and State University, Blacksburg. 2. Neu, H. C., and G. J. Garvey. 1975. Comparative in vitro activity and clinicalipharmacology of ticarcillin and carbenicillin. Antimicrob. Agents Chemother. 8:457-462. 3. Pines, A., G. Khaja, H. Raafat, and K. S. Shreedharan. 1974. Preliminary clinical experience with Ticarcillin (BRL2288) in 101 patients tteated for severe respiratory infections. Chemotherapy 20:3944. 4. Rodriguez, V., G. P. Bodey, N. Horikoshi, J. Inagaki, and K. B. McCredie. 1973. Ticarcillin therapy of infections. Antimicrob. Agents Chemother. 4:427431.
261
5. Simon, C., M. Leuth, and V. Malerczyk. 1974. Ticarcillin. Deutsch. Med. Woschenschr. 99:2460-2464. 6. Steers, E., E. L. Foltz, and B. S. Graves. 1959. An inocula replicating apparatus for routine testing of bacterial susceptibiliiy to antibiotics. Antibiot. Chemother. 9:307-311. 7. Sutter, V. L., and S. M. Finegold. 1975. Susceptibility of anaerobic bacteria to carbenicillin, cefoxitin and related drugs. J. Infect. Dis. 131:417422. 8. Wise, R., and D. S. Reeves. 1974. Clinical and laboratory investigations on Ticarcillin and anti-pseudomonal antibiotic. Chemotherapy 20:45-51.
