Bactericidal Activity of Macromomycin, an Antitumor Antibiotic

Susceptibility testing by the broth dilution method showed that all the gram-positive but only some of the gram-negative bacteria tested were susceptible to the antitumor antibiotic, macromomycin (MCR; NSC 170105). The minimal inhibitory concentration for the susceptible organisms was less than 3 mug/ml. The action of MCR was bactericidal; however, at very high concentrations (50 mug/ml and above) some organisms occasionally escaped death. None of the escaped organisms was resistant to MCR. In combination with other commonly used antibiotics, MCR displayed partial synergy for Pseudomonas aeruginosa (from a minimal inhibitory concentration of >100 to 12.5 mug/ml with 100 mug of chloramphenicol per ml) and for Bacillus pumilus and Staphylococcus aureus (from 1.6 to 0.4 mug/ml and below) with polymyxin B. As with mammalian cells, (125)I-labeled MCR was irreversibly bound to both gram-positive and -negative bacteria. Treatment with trypsin of the (125)I-labeled MCR-exposed cells did not release the bound MCR or reverse its lethal effect. When in solution in a protective buffer at 4 degrees C, MCR was stable for up to 45 days; at 37 degrees C, however, 25% of its bactericidal activity was lost in 72 h. Loss of activity was enhanced 16-fold in the presence of both heated and unheated pooled human sera. Urine had no effect on the activity of MCR.     

Antibiotic Concentrations in Serum, Serum Bactericidal Activity, and Results of Therapy of Streptococcal Endocarditis in Rabbits

The correlation between antibiotic concentrations in serum, serum bactericidal activity, and results of therapy was studied in rabbits with streptococcal endocarditis. Five days of procaine penicillin G reduced bacterial titers to <10 per g in 12 of 14 vegetations in rabbits receiving 75,000 U intramuscularly every 6 h and 10 of 20 in rabbits given 37,500 U. Ten days of 18,750 U every 6 h did not reduce the titers. To test for cure, rabbits were treated with 75,000 U every 6 h for 10 or 20 days and then received no therapy for 7 days. At the end of the 7-day period without therapy, vegetations were sterile in five of five and eight of eight animals, respectively. Rabbits received 37,500 U every 6 h for 5, 10, or 20 days and then no therapy for 7 days, after which vegetations were sterile in one of seven, four of nine, and seven of eight animals, respectively. The median maximal serum bactericidal dilutions at 1 h were 1/16 when 75,000 U of procaine penicillin G was administered, 1/8 to 1/16 with 37,500 U, and 1/4 to 1/8 with 18,750 U. Serum bactericidal activity could not be detected in 50% of the rabbits 6 h after administration of 37,500 U. Cure was related to a median maximal serum bactericidal dilution of at least 1/8 to 1/16 1 h after penicillin administration. A median maximal serum bactericidal dilution of 1/4 to 1/8 resulted in unsuccessful therapy.     

Phagocytic and Bactericidal Properties of Normal Human Monocytes

The bactericidal and phagocytic capacities of monocytes for E. coli, Staphylococcus, Salmonella, and Listeria, and factors that influence these functions were evaluated and compared with those of the polymorphonuclear leukocytes of 30 normal human subjects. Monocytes killed a significantly smaller proportion of each of the bacterial species than did neutrophils from the same individuals. Whereas the neutrophils of all individuals demonstrated the ability to kill significant numbers of the four bacterial species, there was a marked variation in the effect of monocytes of different individuals on the growth curves of these same bacteria. When the bactericidal capacity of an individual's monocytes to more than one species of bacteria was examined in the same experiment, a significant difference in the effect of monocytes on the growth curve of one bacterial species as opposed to another was noted in 4 of 17 subjects. The bactericidal ability of monocytes of single individuals was consistent on different days in 9 of the 11 subjects whose monocytes were examined more than once against the same bacteria.Studies were performed to determine if the lesser bactericidal capability of monocytes was due to a difference in the ability of monocytes and neutrophils to phagocytize or to a difference in the ability of these cells to kill ingested bacteria or both. The results demonstrated that monocytes phagocytize bacteria significantly less well than neutrophils, but the intracellular killing capacity of both cell types is equal. Addition of phenylbutazone to cell suspensions completely inhibited intracellular killing by both monocytes and neutrophils, suggesting the possibility that the bactericidal mechanisms in both cell types might be similar.Monocyte killing of E. coli, Salmonella, and Listeria, but not of Staphylococcus, was significantly diminished in heat-inactivated autologous serum. Neither increasing the concentration of autologous serum from 10% to 25% nor replacement of autologous serum with pooled human serum had any effect on monocyte killing of any of the four bacteria.These studies demonstrate that peripheral blood monocytes are less bactericidal for the four bacterial species than neutrophils, solely because monocytes are less phagocytic. A baseline for further study of factors that influence monocyte function and for study of this cell in selected patient populations is provided.     

Comparison of Serum Bactericidal Activity Among Three Antimicrobial Combinations

Three antimicrobial combinations, ticarcillin plus cephalothin (T+C), ticarcillin plus gentamicin (T+G), and cephalothin plus gentamicin (C+G), were administered to 105 febrile granulocytopenic cancer patients at the Baltimore Cancer Research Center as part of a multi-institutional prospective randomized antibiotic trial. The sera from 32 of these patients (T+C−10 patients, T+G−10 patients, and C+G−12 patients) obtained 1 h post-antibiotic administration were examined for bactericidal activity against 11 strains each of the most common pathogens infecting the granulocytopenic host: Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, and Staphylococcus aureus. Each of the three antibiotic regimens produced a high degree of bactericidal activity in these sera against S. aureus and E. coli. P. aeruginosa was equally, although poorly, killed by sera containing ticarcillin (T+G, T+C), whereas C+G produced no measurable serocidal activity (P < 0.05). Sera with C+G killed K. pneumoniae more effectively than T+G; T+C produced the least killing effect of the three regimens against this organism (P < 0.05). The bactericidal activity of the serum from these 32 patients supplements the overall clinical results of the multi-institutional antibiotic trial and suggests that T+G is a useful initial regimen for empiric therapy of febrile episodes in granulocytopenic cancer patients.     

正常人及脐血清过氧化氢酶活性的参考范围

本文应用分光光度比色法，检测了９７７例从１～＞６０岁不同年龄组的正常人血清以及４７例正常分娩的新生儿脐血清ＣＡＴ活性。调查结果表明，男性５０１例，血清ＣＡＴ活性为：５５．７±１７．９ＫＵ／Ｌ（１５．６～１１８．２）．女性组４７６例，血清ＣＡＴ活性为４９．７±１６．１ＫＵ／Ｌ（１５．２～９８．８）。二者平均值为５２．７±１７．０ＫＵ／Ｍ１５．２～１１８．２）．男性组较女性组血清ＣＡＴ活性高１６．６％（Ｐ＜Ｏ．０１）．１～＞６０岁不同年龄段的正常人血清ＣＡＴ活性有随年龄增长而增加的倾向，各年龄组间血清ＣＡＴ活性经Ｆ检验，Ｐ＞０．０５．新生儿脐血清ＣＡＴ活性的平均值是：１４７．７±５９．９ＫＵ／Ｌ（４２．０～２３６．４），大约为正常人血清ＣＡＴ活性的２．８倍。     

In Vitro Bactericidal Activity of Iclaprim in Human Plasma

This study evaluated the effect of human plasma on the in vitro bactericidal activity of the novel diaminopyrimidine iclaprim against methicillin (meticillin)-susceptible and -resistant Staphylococcus aureus strains. MICs and minimal bactericidal concentrations (MBCs) of iclaprim, with ∼93% protein binding, were similar in the absence and in the presence of 50% human plasma; MICs and MBCs ranged from 0.06 to 0.125 μg/ml. Furthermore, the activity of iclaprim was not affected by plasma, with ≥99.9% reduction in CFU after 5.0 to 7.6 h.Iclaprim is a novel dihydrofolate reductase inhibitor antibiotic that is under development for the treatment of hospitalized patients with severe infections caused by gram-positive pathogens, including Staphylococcus aureus, methicillin (meticillin)-resistant S. aureus (MRSA), and β-hemolytic streptococci (10, 11). Iclaprim has recently completed two pivotal phase III trials for the treatment of complicated skin and skin structure infections, including infections caused by MRSA. It has been shown to exhibit a rapid in vitro bactericidal activity against MRSA and vancomycin-nonsusceptible strains (11).Plasma protein binding is often, though not always, associated with a certain loss in in vitro microbiological activity. Despite the observation that iclaprim is ∼93% plasma protein bound, we have recently reported that the in vitro MIC of iclaprim is not affected by the presence of 50% human plasma (7). In contrast, the microbiological activity of fusidic acid (97 to 98% plasma protein bound) has been reported to be significantly affected by the addition of serum or blood to the test medium (4). The aim of this study was to determine the effect of human plasma on the bactericidal activity of iclaprim against S. aureus in comparison to the activity of teicoplanin and fusidic acid drugs with similar or higher protein binding (>90% and 97 to 98%, respectively) and vancomycin and linezolid with low reported protein binding (55% and 31%, respectively) (2).Ten methicillin-susceptible S. aureus (MSSA) and 10 MRSA strains were tested, including clinical isolates from different countries in Europe and North America and two type strains. MICs against iclaprim, teicoplanin, vancomycin, fusidic acid, and linezolid were determined by broth microdilutions in cation-adjusted Mueller-Hinton broth (CAMHB; Oxoid, Basingstoke, United Kingdom) with and without 50% pooled human plasma (PAA Laboratories GmbH, Pasching, Austria) following the standard CLSI protocol (3). Minimal bactericidal concentrations (MBCs) were determined via plating of aliquots on Mueller-Hinton agar taken from wells with no visual growth at 24 h, according to the CLSI guideline (9). To avoid the impact of potential drug carryover, cells grown with fusidic acid were washed twice with phosphate-buffered saline before plating (6). Washing did not affect the CFU counts. The rates of the bactericidal activities of iclaprim and vancomycin in the presence of 50% human plasma were assessed for one MSSA and four MRSA strains by time-kill methodology (9). The bacteria (∼1 × 10⁶ CFU/ml) were grown in 24-well plates containing 2 ml of CAMHB either with or without 50% human plasma and either containing no antibiotic (control) or containing iclaprim or vancomycin at 4× MIC, and the CFU/ml were determined. Samples were taken at 0, 2, 4, 6, 8, and 24 h, and appropriate dilutions were plated onto Mueller-Hinton agar to determine the CFU. Bactericidal activity was defined as a ≥3-log₁₀ CFU/ml reduction in bacterial density (i.e., ≥99.9% kill) compared with the level in the initial inoculum (9). Bacteria growing in the presence of plasma were sonicated briefly to disaggregate cellular clumps that can form in the presence of human plasma (Branson Sonifier 250; 30 s at 40% duty cycle and 40% capacity), which can result in an underestimation of CFU. The gentle sonication did not affect the viability of the cells.The activity of iclaprim was not affected by the presence of 50% human plasma, with MICs ranging from 0.06 to 0.125 μg/ml with and without 50% human plasma, in agreement with recently published data (7). Moreover, the MBCs of iclaprim were also not affected by the presence of human plasma. The MBC/MIC ratios ranged from 1 to 2 both in CAMHB and in the presence of 50% human plasma (Table ​(Table1).1). Therefore, iclaprim exhibited similar bactericidal activity irrespective of the presence or absence of 50% plasma according to MBC determinations. The activities of vancomycin and linezolid were also generally unaffected by the presence of human plasma, with similar MICs and MBCs in the presence and absence of 50% plasma (Table ​(Table1).1). Vancomycin was bactericidal and the MBC/MIC ratios ranged from 1 to 2, while linezolid was bacteriostatic (Table ​(Table1),1), which is in agreement with reported data (1, 12). The presence of plasma had a minimal effect on the activity of teicoplanin (Table ​(Table1).1). These data are in agreement with Dykhuizen et al., who reported similar MBCs of vancomycin and teicoplanin in the presence of 50% human serum (5). As expected from its high plasma protein binding, MICs for fusidic acid were 16- to 256-fold greater in the presence of plasma, which is in agreement with previously published data (4, 7). MBC ranges were 0.03 to >8 μg/ml in CAMHB and 8 to >128 μg/ml in the presence of 50% human plasma (Table ​(Table11).

TABLE 1.

Antibacterial activities of iclaprim and comparators against 20 isolates of S. aureus^a

Bactericidal Action of an Antibiotic Produced by Myxococcus xanthus

Myxococcus xanthus produced an antibiotic during the end of its exponential growth phase which was capable of inhibiting growth of several gram-positive and gram-negative bacteria. The antibiotic was bactericidal to growing cultures only; chloramphenicol inhibited the bactericidal action of the antibiotic. Upon addition of the antibiotic to Escherichia coli B, deoxyribonucleic acid and ribonucleic acid as well as turbidity of the culture continued to increase even after the viable count decreased; the culture lysed about 60 min after addition of sufficient concentrations of the antibiotic. Spheroplasts could be prepared if the antibiotic was added to a culture growing in the presence of high concentrations of sucrose and MgSO₄. Mutants of M. xanthus FB which are incapable of fruiting body formation or glycerol-induced myxospore formation also produced the antibiotic. A mutant of E. coli resistant to the purified antibiotic was isolated in order to study the role of the antibiotic in the predatory behavior of myxococci.     

Comparison of Broth and Human Serum as the Diluent in the Serum Bactericidal Test

The use of serum rather than broth as the diluent in the serum bactericidal test results in a significant decrease in the test level among patients receiving highly protein-bound semisynthetic penicillins.     

Bactericidal Activity of Cephalosporins in an In Vitro Model Simulating Serum Levels

The bactericidal activity of cefazolin, cephaloridine, and cephalothin in a simulated intramuscular study (500 mg) and a simulated intravenous drip infusion study (2 g/2 h) is reported. In both model systems, the bactericidal activity of cefazolin surpassed that of cephalothin, and there were certain differences between cefazolin and cephaloridine in the simulated intramuscular study when human serum was used as a medium. In a routine reference static system, the drug levels were constant at the simulated peak level of each cephalosporin by both routes. In this system the three cephalosporins were equal in activity. In a third experiment, the effect of drug concentrations and exposure time on bactericidal activity of the cephalosporins was studied. The bactericidal activity of cephaloridine was the strongest of the three drugs when exposure time was 2 h and drug concentration was less than four times the minimal inhibitory concentration. At concentrations above four times the minimum inhibitory concentration, all three cephalosporins were equal in activity when the exposure time was 2 h.     

Chemotactic Factor Inactivator in Normal Human Serum

Normal human serum contains an inactivator of chemotactic factors for neutrophilic leukocytes. The chemotactic factor inactivator (CF-I) remains soluble when serum is fractionated with ammonium sulfate (at 45% saturation), directly and irreversibly inactivates chemotactic factors, and it has a broad spectrum of activity as indicated by its inactivation of the chemotactic fragments of human C3 and C5 (third and fifth components of complement), C, and the bacterial chemotactic factor derived from Escherichia coli. CF-I appears as a biphasic activity according to preparative techniques of sucrose density ultracentrifugation, electrophoresis, and gel filtration. Studies on the interaction of CF-I with the radiotagged C5 chemotactic fragment fail to reveal evidence for irreversible binding as the basis for inactivation. CF-I varies from the anaphylatoxin inactivator in several physical-chemical respects, but evidence does not permit a conclusive statement about the relationship of the two inactivators. CF-I may function as a regulator of inflammatory responses.     

Effects of Cytochalasin B on the Intracellular Bactericidal Activity of Human Neutrophils

Cytochalasin B (CB) is known to have some inhibitory effects on cytokinesis, single-cell movement, bacterial uptake by phagocytes, and many other processes. The effects of CB on intraleukocytic bactericidal activities in human leukocytes were studied, and the results were summarized as follows. (i) CB inhibited the early stage of the intracellular bactericidal activity of human leukocytes against Streptococcus pyogenes D58 (group A). The effect was rapidly eliminated by rinsing the CB solution. (ii) In the late stage of the intracellular bactericidal process, however, CB possessed no effect against S. pyogenes D58 (group A) and Staphylococcus aureus 209P. (iii) CB also inhibited the translocation of myeloperoxidase granules to the phagosomes of human neutrophils.     

Deficient Serum Bactericidal Activity Against Escherichia Coli in Patients with Cirrhosis of the Liver

The serum bactericidal activity (SBA) of cirrhotic patients was compared with that of normal individuals using the release of (51)Cr from radiolabeled Escherichia coli as the assay method. 80% (22/27) of patients were found to have deficient SBA against at least one of three smooth, serum-sensitive test strains of E. coli. Cirrhotic patients were found to have normal levels of serum lysozyme. Although some patients were mildly hypocomplementemic, this abnormality did not correlate with the presence of a bactericidal defect. Bactericidal antibody in normal and cirrhotics' sera was limited to the immunoglobulin (Ig)M class. Purified IgM from patients with deficient SBA against E. coli 0111 had lower concentrations of bactericidal antibody for that E. coli than did IgM from normal sera; the calculated bactericidal activity of total serum IgM was also lower. The bactericidal defect in cirrhotic serum could be completely corrected by either human antiserum to the homologous strain of E. coli or by purified, normal human IgM. However, because higher concentrations of IgM were required to restore normal SBA to a cirrhotic's serum than to agammaglobulinemic serum, there may be an inhibitor of bactericidal antibody in addition to a deficiency of bactericidal IgM antibody to E. coli in the serum of patients with cirrhosis. The bactericidal activity of the alternative complement pathway was also assessed. Sera from cirrhotic patients had no deficit in SBA attributable to the alternative complement pathway. In fact, in some, the activity of the alternative complement pathway was supernormal, compensating in part for the deficit in IgM-mediated SBA.     

Bactericidal Activity and Pharmacology of Cefazolin

The in vitro bactericidal activity of cefazolin was found to be similar to that of cephaloridine and cephalothin but slightly greater than that of cephalexin against a majority of 233 strains of gram-positive and gram-negative organisms. Cefazolin, however, was two- to eightfold more active than the other two drugs against Escherichia coli and klebsiella. The mean peak concentrations in the serum in 10 normal subjects 1 h after intramuscular injections of 1,000, 500, and 250 mg of cefazolin were 38.8, 18.6, and 12.2 mug/ml, respectively. The antibiotic could still be detected at 8 h. Peak values for a given dose of cephaloridine were comparable. However, blood levels of cefazolin were regularly higher than those of cephaloridine over the first 8 h. The mean half-life of cefazolin in the serum was 2 h, whereas that of cephaloridine was 1.4 h. The degree of serum protein-binding was strikingly higher for cefazolin (81%) than for cephaloridine (24%), suggesting that the antibacterial activity of the former in serum might be less than that of cephaloridine after equal doses. This proved to be the case when the bacterial activity of blood drawn 1, 4, and 8 h after injection of the two drugs was examined.     

Bactericidal Activity of the Human Skin Fatty Acid cis-6-Hexadecanoic Acid on Staphylococcus aureus

Human skin fatty acids are a potent aspect of our innate defenses, giving surface protection against potentially invasive organisms. They provide an important parameter in determining the ecology of the skin microflora, and alterations can lead to increased colonization by pathogens such as Staphylococcus aureus. Harnessing skin fatty acids may also give a new avenue of exploration in the generation of control measures against drug-resistant organisms. Despite their importance, the mechanism(s) whereby skin fatty acids kill bacteria has remained largely elusive. Here, we describe an analysis of the bactericidal effects of the major human skin fatty acid cis-6-hexadecenoic acid (C6H) on the human commensal and pathogen S. aureus. Several C6H concentration-dependent mechanisms were found. At high concentrations, C6H swiftly kills cells associated with a general loss of membrane integrity. However, C6H still kills at lower concentrations, acting through disruption of the proton motive force, an increase in membrane fluidity, and its effects on electron transfer. The design of analogues with altered bactericidal effects has begun to determine the structural constraints on activity and paves the way for the rational design of new antistaphylococcal agents.     

Bactericidal Activity of Vancomycin in Cerebrospinal Fluid

Intraventricular application of vancomycin is an effective therapeutic regimen for the treatment of shunt-associated staphylococcal ventriculitis. We examined the in vitro activity of vancomycin at high concentrations against Staphylococcus aureus ATCC 25923 and Staphylococcus epidermidis ATCC 12228 in human cerebrospinal fluid samples. Time-kill curves revealed equal efficacies for concentrations of 10, 100, and 300 microg/ml, and incubation times of 24 to 48 h were needed to achieve a 3 log(10) reduction of viable bacteria. A concentration of 5 microg/ml showed a slightly lower activity, but this difference was not significant. In an infant who was successfully treated for shunt-associated ventriculitis due to S. epidermidis by once-daily local administration of vancomycin (3 mg for 2 days and 5 mg for 4 days [0. 5 to 0.8 mg/kg of body weight]) the in vivo kill kinetics were similar to those for the in vitro results. These results support time-dose regimens that provide trough vancomycin levels of 5 to 10 microg/ml.     

人脐血清造血刺激活性的检测

通过半固体培养法(甲基纤维素法和琼脂培养法)和液体长期培养等方法,观察了脐血清和成人血清对正常小鼠骨髓细胞的影响。结果表明:①脐血清EPO活性高于成人血清,红系祖细胞产率分别为23.2±5.4/1×10~5细胞和3.0±1.3/1×10~5细胞,差异非常显著;对PHA刺激的小鼠脾细胞培养液的活性,脐血清抑制作用较低;②以脐血清作为外源性刺激因子,可形成粒单系祖细胞21.7±16.8集落和丛/1×10~5细胞,用成人血清仅见个别小丛1.1±1.2丛/1×10~5细胞;③脐血清可促进成纤维系祖细胞(CFU-F)增殖与分化,其产率为36.0±29.7/1×10~6细胞,明显高于成人血清。提示:脐血清不仅含有丰富的红系、粒单系、巨核系祖细胞,而且造血活性也高于成人血清,因而脐血输注和移植将有广阔前景。     

Serum Bactericidal Activity of Moxalactam and Cefotaxime With and Without Tobramycin Against Pseudomonas aeruginosa and Staphylococcus aureus

Serum bactericidal activity was determined against 10 strains each of Pseudomonas aeruginosa and Staphylococcus aureus in serum from volunteers 1 and 6 h after intravenous infusion of cefotaxime, tobramycin, and the combination; or of moxalactam, tobramycin, and the combination. High serum bactericidal activity against P. aeruginosa was found significantly more frequently with moxalactam plus tobramycin than with cefotaxime, moxalactam, and tobramycin alone or with cefotaxime plus tobramycin.     

Bactericidal versus Bacteriostatic Antibiotic Therapy of Experimental Pneumococcal Meningitis in Rabbits

A rabbit model of pneumococcal meningitis was used to examine the importance of bactericidal vs. bacteriostatic antimicrobial agents in the therapy of meningitis 112 animals were infected with one of two strains of type III Streptococcus pneumoniae. Both strains were exquisitely sensitive to ampicillin, minimum inhibitory concentration (MIC)/minimum bactericidal concentration (MBC)<0.125 μg/ml. The activity of chloramphenicol against the two strains varied: strain₁—MIC 2 μg/ml, MBC 16 μg/ml; strain₂—MIC 1 μg/ml, MBC 2 μg/ml. Animals were treated with either ampicillin or chloramphenicol in dosages that achieved a peak bactericidal effect in cerebrospinal fluid (CSF) for ampicillin against both strains. Two different dosages were used for chloramphenicol. The first dosage achieved a peak CSF concentration of 4.4±1.1 μg/ml that produced a bacteriostatic effect against strain₁ and bactericidal effect against strain₂. The second dosage achieved a bactericidal effect against both strains (mean peak CSF concentration 30.0 μg/ml). All animals were treated intramuscularly three times a day for 5 d. CSF was sampled daily and 3 d after discontinuation of therapy for quantitative bacterial cultures. Results demonstrate that only antimicrobial therapy that achieved a bactericidal effect in CSF was associated with cure. Over 90% of animals treated with one of the bactericidal regimens (i.e., animals in which the bacterial counts in CSF dropped >5 log₁₀ colony-forming units [cfu]/ ml after 48 h) had sterile CSF after 5 d of treatment. On the other hand, the regimen that achieved bacteriostatic concentrations (CSF drug concentrations between the MIC and MBC) produced a drop of 2.4 log₁₀ cfu/ml by 48 h; however, none of the animals that survived had sterile CSF after 5 d. These studies clearly demonstrate in a strictly controlled manner that maximally effective antimicrobial therapy of experimental pneumococcal meningitis depends on achieving a bactericidal effect in CSF.