THE JOURNAL OF INFECTIOUS DISEASES. VOL. 136, NO.3. SEPTEMBER 1977 © 1977 by the University of Chicago. All rights reserved.
MAJOR ARTICLES Experimental Endocarditis Due to Pseudomonas aeruginosa. II. Therapy with Carbenicillin and Gentamicin From the Division of Infectious Diseases, Department of Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia, and Ann Arbor Veterans Administration Hospital, Ann Arbor, Michigan
Gordon Archer and F. Robert Fekety, Jr.
Antibiotic therapy of endocarditis caused by Pseudomonas aeruginosa has been curative in only one-third of drug addicts with tricuspid valve infections and in even fewer patients with infected left-sided or prosthetic valves [1-5]. Surgical removal of infected tissues is usually required for cure [6]. To study systematically antibiotic regimens that may be better, we devised an experimental model of left-sided P. aeruginosa endocarditis in rabbits [7]. This report describes the results of our studies of the Received for publication January 29, 1976, and in revised form January 24,1977. This work was presented in part at the 14th Interscience Conference on Antimicrobial Agents and Chemotherapy. San Francisco, California, on September II, 1974. This study was supported by research funds from the Veterans Administration Hospital. an institutional grant from Virginia Commonwealth University, and a grant from the Mi 0.05). However, in statistical analysis, combination therapy was significantly better than therapy with gentamicin alone (P < 0.05) if those two rabbits with renal failure and sustained high serum bactericidal activity in the gentamicin group were excluded. Levels of gentamicin in serum in these two rabbits were probably not comparable to levels in other animals given the same dose of gentamicin. Only two (8.3%) of 24 rabbits treated with the combination had positive blood culture results during therapy compared with 14 (50%) of 28 treated with 5 mg of gentamicin/kg (P < 0.005) and 14 (54%) of 26 treated with 400 mg of carbenicillin/kg (P < 0.005). Only gentamicin in a dose of 7.5 mg/kg was comparable to the combination of gentamicin and carbenicillin in ability to sterilize the blood. None of eight cultures of renal cortical abscesses in animals given the combination regimen contained bacteria. Six animals treated with the combination for 11 days were followed after treatment; none relapsed during the two-week observation period. Five had sterile vegetations at sacrifice, whereas one had 105 . 0 cfu of P. aeruginosa / g cultured from small vegetations. Serum bactericidal activity. Table 3 summarizes the bactericidal activity of sera from treated animals against the challenge organism. Sera
Table 3. Bactericidal titers in sera of rabbits with endocarditis due to Pseudomonas aeruginosa measured at various intervals after injection of antibiotics. Titer at indicated time after injection
Treatment group (no. of animals evaluated) Untreated controls (5) Gentamicin,5 mg/kg (14)* Carbenicillin, 400 mg/kg (14) Combination of 5 mg of gentamicin/kg and 400 mg of carbenicillin/kg (17) Gentamicin, 1.5 mg/kg (17)
1 hr
o
4 hr
2 (2-4)
o o
8 (4-16)
o
16 (8-32) 32 (16-64)
2 (0-4) 2 (0-4)
NOTE. Data are median reciprocal titers with ranges in parentheses. All values at 7 hr were O. *Titers for two rabbits with renal failure are not included.
Downloaded from http://jid.oxfordjournals.org/ at Cambridge University on August 31, 2015
died after discontinuation of gentamlcm; one died one week and three died two weeks after the end of therapy. Culture of vegetations from these rabbits yielded 107 . 5 ± 101.0 cfu of P. aeruginosa / g of vegetation. Of two survivors with no clinical evidence of endocarditis that were sacrificed two weeks after therapy had been discontinued, one had no bacteria and the other had 106 .0 cfu/g cultured from vegetations. Carbenicillin therapy. Ten rabbits were treated with carbenicillin for six days and 13 for two weeks. Levels of carbenicillin detected in serum after a 400-mg/kg im dose are shown in table 1. Levels found I hr after injection are similar to those recommended in humans who are being treated for infections due to P. aeruginosa. Although the mortality rate was low during the first six days of treatment with carbenicillin, it increased markedly during the second week. No vegetations were sterilized after either six or 14 days of therapy, and high bacterial densities were found in vegetations (table 2). Ten of 13 rabbits treated for two weeks developed positive blood culture results, with 102-104 cfu of P. aeruginosajml of blood. Cultures of renal cortical abscesses from five of eight rabbits yielded P. aeruginosa) with a mean of 106 .8 ± 10°·8 cfu/g of tissue. Four rabbits that were alive after two weeks of therapy were followed for detection of relapses, and all four died within a week of discontinuation of antibiotics. Cultures of their vegetations yielded 108.8 ± 10°. 8 cfu of P. aeruginosa / g of tissue. Combination of carbenicillin and gentamicin therapy. Ten rabbits were treated with the combination of 400 mg of carbenicillin/kg and 5 mg of gentamicin/kg for six days, and 14 animals were treated with this antibiotic regimen for two weeks. Only three animals died during the course of therapy. Although no vegetations were sterilized after six days of combination therapy, the number of P. aeruginosa in vegetations (table 2) was significantly less than that in vegetations of rabbits treated with either drug alone (P < 0.05). Tissue adherent to the catheter and material obtained from valves or endocardium contained equal quantities of bacteria. The number of animals with vegetations sterilized by two weeks of combination therapy was significantly greater than the number with vegeta-
332
of the other seven rabbits survlvmg after two weeks of treatment with carbenicillin alone, including four animals that relapsed, had cultures that yielded P. aeruginosa with increased resistance to carbenicillin. Vegetations from some treated and untreated rabbits yielded P. aeruginosa that displayed marked colonial variation. Both large- and smallcolony variants were selected and retested for growth in broth, biochemical characteristics, and antibiotic susceptibility. Approximately half of the small-colony variants reverted to normal colonial morphology after incubation in broth, whereas the other half retained the small-colony characteristic after repeated subculture. There were otherwise no differences in the large- and small-colony variants with regard to antibiotic susceptibility, pigment production, biochemical characteristics, or immunotype. Antibiotic toxicity. Serum was obtained from 43 rabbits before and after gentamicin therapy for determination of levels of creatinine; five (12%) had abnormal levels after treatment. In three rabbits changes in levels before and after treatment were minor, i.e., from 0.8, 0.9, and 1.2 mg/ dl before treatment to 1.4, 1.4, and 3.5 mg/ dl, respectively, after treatment. Serum bactericidal titers in these three rabbi ts were similar to those in other rabbits in their treatment group, and microscopic renal pathology was no different than that seen in untreated rabbits or in treated rabbits with stable levels of creatinine. This pathology was presumably attributable to endocarditis and consisted of focal areas of cortical inflammation and infarction necrosis with normal glomeruli in uninfarcted areas and no evidence of diffuse tubular abnormalities. In two rabbits, however, levels of creatinine of 8.4 and 14.6 mg/ dl, respectively, after treatment were markedly elevated from levels of 1.2 and 0.7 mg/ell, respectively, before treatment. Serum bactericidal activity in these rabbits on day 6 of therapy was similar to that seen in other animals treated with this regimen of gentamicin therapy (table 3). By day 12 bactericidal titers in serum had increased to a peak of 1: 128 and were still 1:32 before the next dose of drug. The kidneys from these animals showed microscopic evidence of diffuse tubular necrosis. All of the elevated levels of creatinine were in rabbits treated with gentamicin alone. One
Downloaded from http://jid.oxfordjournals.org/ at Cambridge University on August 31, 2015
ranged from pH 7.2 to pH 7.6. There was no difference in the mean activity of sera obtained early in treatment vs. that in sera obtained later, with the exception of the two animals that developed marked renal insufficiency during the course of therapy (see below). The highest peak activities were reported with the higher dose of gentamicin and the combination of the lower dose of gentamicin and carbenicillin. These regimens were also the only ones that produced detectable bactericidal activity in serum 4 hr after treatment. Peak bactericidal activity in serum directly correlated with sterilization of vegeta· tions. After treatment for two weeks, nine (60%) of 15 rabbits with peak bactericidal titers in broth of ~l: 16 had sterile vegetations, whereas only two (13%) of 15 rabbits with titers of
MAJOR ARTICLES Experimental Endocarditis Due to Pseudomonas aeruginosa. II. Therapy with Carbenicillin and Gentamicin From the Division of Infectious Diseases, Department of Medicine, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia, and Ann Arbor Veterans Administration Hospital, Ann Arbor, Michigan
Gordon Archer and F. Robert Fekety, Jr.
Antibiotic therapy of endocarditis caused by Pseudomonas aeruginosa has been curative in only one-third of drug addicts with tricuspid valve infections and in even fewer patients with infected left-sided or prosthetic valves [1-5]. Surgical removal of infected tissues is usually required for cure [6]. To study systematically antibiotic regimens that may be better, we devised an experimental model of left-sided P. aeruginosa endocarditis in rabbits [7]. This report describes the results of our studies of the Received for publication January 29, 1976, and in revised form January 24,1977. This work was presented in part at the 14th Interscience Conference on Antimicrobial Agents and Chemotherapy. San Francisco, California, on September II, 1974. This study was supported by research funds from the Veterans Administration Hospital. an institutional grant from Virginia Commonwealth University, and a grant from the Mi 0.05). However, in statistical analysis, combination therapy was significantly better than therapy with gentamicin alone (P < 0.05) if those two rabbits with renal failure and sustained high serum bactericidal activity in the gentamicin group were excluded. Levels of gentamicin in serum in these two rabbits were probably not comparable to levels in other animals given the same dose of gentamicin. Only two (8.3%) of 24 rabbits treated with the combination had positive blood culture results during therapy compared with 14 (50%) of 28 treated with 5 mg of gentamicin/kg (P < 0.005) and 14 (54%) of 26 treated with 400 mg of carbenicillin/kg (P < 0.005). Only gentamicin in a dose of 7.5 mg/kg was comparable to the combination of gentamicin and carbenicillin in ability to sterilize the blood. None of eight cultures of renal cortical abscesses in animals given the combination regimen contained bacteria. Six animals treated with the combination for 11 days were followed after treatment; none relapsed during the two-week observation period. Five had sterile vegetations at sacrifice, whereas one had 105 . 0 cfu of P. aeruginosa / g cultured from small vegetations. Serum bactericidal activity. Table 3 summarizes the bactericidal activity of sera from treated animals against the challenge organism. Sera
Table 3. Bactericidal titers in sera of rabbits with endocarditis due to Pseudomonas aeruginosa measured at various intervals after injection of antibiotics. Titer at indicated time after injection
Treatment group (no. of animals evaluated) Untreated controls (5) Gentamicin,5 mg/kg (14)* Carbenicillin, 400 mg/kg (14) Combination of 5 mg of gentamicin/kg and 400 mg of carbenicillin/kg (17) Gentamicin, 1.5 mg/kg (17)
1 hr
o
4 hr
2 (2-4)
o o
8 (4-16)
o
16 (8-32) 32 (16-64)
2 (0-4) 2 (0-4)
NOTE. Data are median reciprocal titers with ranges in parentheses. All values at 7 hr were O. *Titers for two rabbits with renal failure are not included.
Downloaded from http://jid.oxfordjournals.org/ at Cambridge University on August 31, 2015
died after discontinuation of gentamlcm; one died one week and three died two weeks after the end of therapy. Culture of vegetations from these rabbits yielded 107 . 5 ± 101.0 cfu of P. aeruginosa / g of vegetation. Of two survivors with no clinical evidence of endocarditis that were sacrificed two weeks after therapy had been discontinued, one had no bacteria and the other had 106 .0 cfu/g cultured from vegetations. Carbenicillin therapy. Ten rabbits were treated with carbenicillin for six days and 13 for two weeks. Levels of carbenicillin detected in serum after a 400-mg/kg im dose are shown in table 1. Levels found I hr after injection are similar to those recommended in humans who are being treated for infections due to P. aeruginosa. Although the mortality rate was low during the first six days of treatment with carbenicillin, it increased markedly during the second week. No vegetations were sterilized after either six or 14 days of therapy, and high bacterial densities were found in vegetations (table 2). Ten of 13 rabbits treated for two weeks developed positive blood culture results, with 102-104 cfu of P. aeruginosajml of blood. Cultures of renal cortical abscesses from five of eight rabbits yielded P. aeruginosa) with a mean of 106 .8 ± 10°·8 cfu/g of tissue. Four rabbits that were alive after two weeks of therapy were followed for detection of relapses, and all four died within a week of discontinuation of antibiotics. Cultures of their vegetations yielded 108.8 ± 10°. 8 cfu of P. aeruginosa / g of tissue. Combination of carbenicillin and gentamicin therapy. Ten rabbits were treated with the combination of 400 mg of carbenicillin/kg and 5 mg of gentamicin/kg for six days, and 14 animals were treated with this antibiotic regimen for two weeks. Only three animals died during the course of therapy. Although no vegetations were sterilized after six days of combination therapy, the number of P. aeruginosa in vegetations (table 2) was significantly less than that in vegetations of rabbits treated with either drug alone (P < 0.05). Tissue adherent to the catheter and material obtained from valves or endocardium contained equal quantities of bacteria. The number of animals with vegetations sterilized by two weeks of combination therapy was significantly greater than the number with vegeta-
332
of the other seven rabbits survlvmg after two weeks of treatment with carbenicillin alone, including four animals that relapsed, had cultures that yielded P. aeruginosa with increased resistance to carbenicillin. Vegetations from some treated and untreated rabbits yielded P. aeruginosa that displayed marked colonial variation. Both large- and smallcolony variants were selected and retested for growth in broth, biochemical characteristics, and antibiotic susceptibility. Approximately half of the small-colony variants reverted to normal colonial morphology after incubation in broth, whereas the other half retained the small-colony characteristic after repeated subculture. There were otherwise no differences in the large- and small-colony variants with regard to antibiotic susceptibility, pigment production, biochemical characteristics, or immunotype. Antibiotic toxicity. Serum was obtained from 43 rabbits before and after gentamicin therapy for determination of levels of creatinine; five (12%) had abnormal levels after treatment. In three rabbits changes in levels before and after treatment were minor, i.e., from 0.8, 0.9, and 1.2 mg/ dl before treatment to 1.4, 1.4, and 3.5 mg/ dl, respectively, after treatment. Serum bactericidal titers in these three rabbi ts were similar to those in other rabbits in their treatment group, and microscopic renal pathology was no different than that seen in untreated rabbits or in treated rabbits with stable levels of creatinine. This pathology was presumably attributable to endocarditis and consisted of focal areas of cortical inflammation and infarction necrosis with normal glomeruli in uninfarcted areas and no evidence of diffuse tubular abnormalities. In two rabbits, however, levels of creatinine of 8.4 and 14.6 mg/ dl, respectively, after treatment were markedly elevated from levels of 1.2 and 0.7 mg/ell, respectively, before treatment. Serum bactericidal activity in these rabbits on day 6 of therapy was similar to that seen in other animals treated with this regimen of gentamicin therapy (table 3). By day 12 bactericidal titers in serum had increased to a peak of 1: 128 and were still 1:32 before the next dose of drug. The kidneys from these animals showed microscopic evidence of diffuse tubular necrosis. All of the elevated levels of creatinine were in rabbits treated with gentamicin alone. One
Downloaded from http://jid.oxfordjournals.org/ at Cambridge University on August 31, 2015
ranged from pH 7.2 to pH 7.6. There was no difference in the mean activity of sera obtained early in treatment vs. that in sera obtained later, with the exception of the two animals that developed marked renal insufficiency during the course of therapy (see below). The highest peak activities were reported with the higher dose of gentamicin and the combination of the lower dose of gentamicin and carbenicillin. These regimens were also the only ones that produced detectable bactericidal activity in serum 4 hr after treatment. Peak bactericidal activity in serum directly correlated with sterilization of vegeta· tions. After treatment for two weeks, nine (60%) of 15 rabbits with peak bactericidal titers in broth of ~l: 16 had sterile vegetations, whereas only two (13%) of 15 rabbits with titers of
