Effect of Penicillin on the Adherence of Streptococcus sanguis In Vitro and in the Rabbit Model of Endocarditis

The effect of penicillin treatment of Streptococcus sanguis in vitro, on subsequent bacterial density in the bloodstream and on cardiac valves in the rabbit model of endocarditis was studied. As experimental tools for this study, isogenic pairs of S. sanguis differing in resistance to streptomycin or rifampin were prepared by genetic transformation. Rabbits with traumatized heart valves received an intravenous inoculation of penicillin treated (1 μg/ml) and untreated S. sanguis, each marked by resistance to either streptomycin or rifampin. The number of penicillin-treated and untreated bacteria attached to the valvular surfaces was determined by differential counting on streptomycin or rifampin containing media. Penicillin pretreatment reduced cardiac valve colonization 5 min after inoculation (“adherence ratio” × 10⁸ was 4.11 for the control and 3.66 for the penicillin-treated bacteria, P < 0.001). The results were not due to differences in serum killing or bacterial densities in the bloodstream. There was no difference in valvular bacterial densities 24 h after bacterial inoculation (adherence ratio × 10⁸, 7.26 untreated vs. 6.34 penicillin-pretreated, P > 0.10).     

Tolerant Response of Streptococcus sanguis to Beta-Lactams and Other Cell Wall Inhibitors

In contrast to group A streptococci or Streptococcus pneumoniae, cells of Streptococcus sanguis (group H) do not exhibit the irreversible effects of penicillin treatment, such as loss of viability or lysis. On the other hand, the same bacteria show typical effects of penicillin, such as morphological alterations, reduction in the rate of cell wall synthesis, and secretion of murein and lipoteichoic acid polymers into the medium. A novel effect of cell wall inhibitors was also noted: treatment with beta-lactams or with fosfomycin, d-cycloserine, or beta-halogeno-d-alanine caused the release of substantial amounts of glycerol lipids into the growth medium. The antibiotic “tolerance” of S. sanguis is interpreted in terms of the hypothesis that the activity of bacterial murein hydrolases is essential for the irreversible effects of cell wall inhibitors.     

Antimicrobial Therapy of Experimental Enterococcal Endocarditis

The successful therapy of enterococcal endocarditis requires prolonged administration of synergistic antibiotic combinations. Controversy has arisen regarding optimal therapy (i) when the organism possesses high-level streptomycin resistance, and (ii) when the patient is allergic to penicillin. This study examines these questions in vitro and in a rabbit model of enterococcal endocarditis. The combination of penicillin with either streptomycin or gentamicin increased the rate of bacterial killing in vitro and in vivo when compared with penicillin alone (P < 0.05) when the test strain was relatively susceptible to streptomycin (minimal inhibitory concentration, 128 mug/ml). Only the combination of penicillin and gentamicin was consistently more effective than penicillin alone (P < 0.01) when the test strain was highly resistant to streptomycin (minimal inhibitory concentration > 150,000 mug/ml). The combination of vancomycin and streptomycin was more rapidly bactericidal than vancomycin alone in vitro and in the animal model against the streptomycin-susceptible strain (P < 0.01). The relative rate of in vitro bacterial killing by various antibiotics and combinations was predictive of the efficacy of these drugs in eradicating enterococci from cardiac vegetation in experimental endocarditis.     

Experimental endocarditis caused by Streptococcus sanguis: single and combined antibiotic therapy.

The effectiveness of penicillin G, fosfomycin, and cefoxitin alone and in combination was studied in vitro and in the treatment of left-sided Streptococcus sanguis endocarditis in rabbits. In vitro, the combinations penicillin G plus fosfomycin, penicillin G plus cefoxitin, and fosfomycin plus cefoxitin were synergistic or partially synergistic for S sanguis. Therapy with the combinations was more effective in eradicating the species from cardiac vegetations that was that with each antibiotic used alone.     

Antibiotic Therapy of Experimental Staphylococcus epidermidis Endocarditis

Endocarditis was produced in rabbits with a methicillin-resistant Staphylococcus epidermidis isolate. Subpopulations resistant to other semisynthetic penicillins and cephalosporins were detected in the isolate. Their presence was probably responsible for the increase in minimum bactericidal concentrations and minimum inhibitory concentrations when tests with high inocula, rather than low inocula were pursued. Rabbits were treated for either 2 or 7 days with nafcillin, cephalothin, cefamandole, vancomycin, rifampin, or gentamicin. Spontaneous death was uncommon in either controls (84% survival) or treated animals (80 to 94% survival). There was no significant difference in the number of bacteria in vegetations of rabbits treated for 7 days with cephalothin, cefamandole, nafcillin, or no antibiotic (control). There was a significant reduction in total bacteria in vegetations of rabbits given vancomycin, gentamicin, or rifampin for 7 days as compared with cephalothin, cefamandole, nafcillin or control. Gentamicin or rifampin sterilized significantly more vegetations after 7 days than cephalothin, cefamandole, nafcillin, or control; rifampin was more effective in sterilizing vegetations than either gentamycin or vancomycin after 2 days. Mutants resistant to 10 μg of rifampin per ml comprised the total bacterial population cultured from vegetations of 2 of 17 rabbits treated with this antibiotic for 7 days; there was no change in the susceptibility of vegetation isolates to other antibiotics. Rifampin, vancomycin, or gentamicin may prove to be more effective in humans than cephalosporins or semisynthetic penicillins in the treatment of methicillin-resistant S. epidermidis endocarditis.     

Infection caused by vancomycin-resistant Streptococcus sanguis II

A patient with bacteremia caused by vancomycin-resistant Streptococcus sanguis II is presented. This rare occurrence suggests that vancomycin may not be a completely reliable antibiotic in the treatment of infections due to viridans species of the genus Streptococcus. Gram-positive isolates from blood and otherwise sterile body fluids should be tested for susceptibility to vancomycin.     

Effects of Low Penicillin Concentrations on Cell Morphology and on Peptidoglycan and Protein Synthesis in a Tolerant Streptococcus Strain

Rates of protein and peptidoglycan synthesis were determined by pulse-labeling techniques before and after treatment of exponentially growing cultures of Streptococcus mutans FA-1 with a number of concentrations of penicillin G (0.05, 0.1, 0.3, and 0.4 μg/ml). These penicillin concentrations were all less than that required to saturate the specific penicillin-binding sites present on the surface of this organism (0.5 μg/ml), but were all greater than and, in fact, were multiples of the minimum inhibitory concentration (0.02 μg/ml). Low concentrations of penicillin G (2.5× the minimum inhibitory concentration) immediately halted the exponential increase in the rate of peptidoglycan synthesis normally expected as the result of cell multiplication, but allowed the rate of peptidoglycan synthesis occurring at the time of penicillin addition to be maintained for almost 1 h. An increased penicillin concentration (5× the minimum inhibitory concentration) allowed the rate of peptidoglycan synthesis occurring at the time of penicillin addition to be maintained for a shorter length of time (~0.67 h). Still greater penicillin concentrations caused an immediate inhibition of the peptidoglycan synthetic rate. The effect of penicillin on the rate of protein synthesis was similar, although less pronounced. Samples were taken for scanning electron microscopy immediately before and after 3 h of treatment with a low (2.5× the minimum inhibitory concentration) concentration of penicillin. The surface areas and volumes of the cells in these samples were calculated from the electron micrographs by using computer reconstruction techniques. From the frequency distributions of surface area, the plots of surface area to volume ratio as a function of surface area, and the pulse-labeling data mentioned previously, low, growth-inhibitory concentrations (2.5× the minimum inhibitory concentration) of penicillin are proposed (i) to inhibit the constriction of the division septum, (ii) to prevent the establishment or maturation of new envelope growth sites, and (iii) to have no immediate effects on the synthesis of cell wall peptidoglycan already in progress at the time of penicillin addition.     

Molecular Characterization of Multidrug Resistance in Streptococcus mitis

Antimicrobial resistance was characterized for 14 strains of Streptococcus mitis. HinfI restriction fragment length mapping of gyrA PCR amplicons from three ciprofloxacin-resistant isolates correlated with mutations associated with such resistance in other organisms. By using PCR, seven erythromycin-resistant strains were found to possess either the mef or ermB gene. Hybridization revealed tet(M) in seven tetracycline-resistant isolates.     

Activity of LY333328 in Experimental Meningitis Caused by a Streptococcus pneumoniae Strain Susceptible to Penicillin

In a rabbit model of Streptococcus pneumoniae meningitis single doses of 10 and 2.5 mg of the glycopeptide LY333328 per kg of body weight reduced bacterial titers in cerebrospinal fluid (CSF) almost as rapidly as ceftriaxone at 10 mg/kg/h (changes in log CFU, -0.29 +/- 0.21 and -0.26 +/- 0.22 versus -0.34 +/- 0.15/ml/h). A dose of 1 mg/kg was bacteriostatic (change in log CFU, 0.01 +/- 0.11/ml/h). In two animals receiving LY333328 at a dose of 40 mg/kg the bacterial titers were reduced by 0.54 and 0.51 log CFU/ml/h. The penetration of CSF by LY333328 was 1 to 5%. The concentrations of lipoteichoic and teichoic acids in CSF and neuronal damage were similar in ceftriaxone- and LY333328-treated animals.     

Myocardial depression and nephrotic syndrome in Streptococcus sanguis endocarditis

Nephrotic syndrome is a most unusual complication of infective endocarditis. We report a case of Streptococcus sanguis endocarditis of the tricuspid valve in a patient with a small congenital ventricular septal defect who developed this complication. Severe myocardial depression also occurred and the clinical behaviour and subsequent recovery of this closely paralleled that of the nephrotic syndrome. These unusual associations, their possible interactions and pathogeneses are discussed.     

Sanguicin, a bacteriocin of oral Streptococcus sanguis.

Streptococcus sanguis strain N-2 was found to produce a bacteriocin (sanguicin) which accumulates intracellularly. It was purified by sequential procedures about 98-fold with a recovery of 37% and appeared to be homogeneous on gel electrophoresis. Sanguicin was heat labile and was destroyed by digestion with pronase. The growth of several species of oral indigenous microorganisms was inhibited by sanguicin, of which Bacteriodes melaninogenicus was most susceptible. Sanguicin acted on susceptible cells as a bacteriostatic agent.     

Antibiotic Therapy of Subacute Bacterial Endocarditis

The incidence of frigidity in women is much higher than most physicians realize. The treatment of this affliction is both prophylactic and curative. Most women with true frigidity must be treated by formal psychiatry, but many can be helped considerably by an understanding and sympathetic physician.     

Single and Combination Antibiotic Therapy of Staphylococcus aureus Experimental Endocarditis: Emergence of Gentamicin-Resistant Mutants

The efficacy of nafcillin and gentamicin used alone and in combination at doses giving serum concentrations comparable to those achieved in patients was studied in rabbits with experimental Staphylococcus aureus endocarditis. The organism used was a penicillinase-producing, methicillin-susceptible, clinical isolate. The addition of gentamicin to nafcillin significantly increased the rate of killing of organisms in valvular vegetations, compared to the effect of nafcillin alone. Gentamicin alone delayed mortality but was not effective in reducing the bacterial populations of the vegetations. Bacteremia persisted in the animals treated with gentamicin alone, in contrast to the groups treated with nafcillin or the combination. Selection of a subpopulation of aminoglycoside-resistant small-colony variants occurred in animals treated with gentamicin alone. This variant was subsequently employed in the rabbit model and produced endocarditis, metastatic infection, and bacteremia comparable to those caused by the parent strain. Animals with infection produced by the variant died later than animals infected by the parent strain. Nafcillin was equally effective in reducing the population of both parent and variant strains in vitro and in therapy of the infected animals. Population studies showed the variant to be a mutant emerging at a rate of 1.9 x 10(-7). It was shown to differ from the parent strain in coagulase and hemolysin production, colonial morphology, and aminoglycoside susceptibility, but was similar by light and electron microscopy and in phage type, pigmentation of colonies, deoxyribonuclease production, mannitol fermentation, and growth rate.     

Interactions between Neisseria sicca and viridin B, a bacteriocin produced by Streptococcus mitis.

Viridin B, a bacteriocin produced by Streptococcus mitis (mitior), is bactericidal to Neisseria sicca. Oxygen consumption by actively growing N. sicca cultures ceased immediately upon exposure to viridin B. Adenosine triphosphate production was slightly enhanced within 1 h of exposure to the bacteriocin but was subsequently repressed. The uptake and incorporation of glucose was prevented in the presence of viridin B. The bacteriocin also blocked uptake of an amino acid mixture in chloramphenicol-pretreated cells. Pretreatment or concomitant treatment with a variety of antibiotics known to inhibit specific synthetic pathways did not alter the inhibition of macromolecular synthesis produced by the bacteriocin. Although viridin B blocks protein and nucleic acid syntheses, no degradation of such macromolecules was observed. The inhibitory effects of viridin B on macromolecular synthesis and on viability required the presence of sufficient nutrients to allow active metabolism of N. sicca. The bacteriocin did not inhibit viability or macromolecular synthesis in anaerobically incubated N. sicca. Thus, active, oxidative metabolism by N. sicca cells is essential for viridin B action. A model for viridin B action is proposed.     

Effect of Elevated Atmospheric Pressure on Penicillin Binding by Staphylococcus aureus and Streptococcus pyogenes

A gas pressure of 68 atm, elicited by helium-oxygen gas mixtures, reduced the susceptibility to penicillin of Staphylococcus aureus but not of Streptococcus pyogenes. The elevated pressure also caused a reduction in the binding of (14)C-penicillin to S. aureus, but not to S. pyogenes. When these studies were extended to glycine incorporation, it was shown that, even without penicillin, pressurization reduced glycine incorporation into the cell wall of S. aureus. Incorporation into other cellular components was not altered by pressurization. Cells grown in a pressurized environment were slightly more susceptible than those grown at 1 atm to rapid change in osmotic pressure. In the presence of penicillin, glycine incorporation into the cell wall was reduced to the same low level at 68 atm and at 1 atm. These results suggest that pressurization renders S. aureus less susceptible to penicillin because it reduces the enzymatic activity of the binding component on the cell, a penicillin-sensitive transpeptidase.     

Lethal Effect of a Heterologous Murein Hydrolase on Penicillin-Treated Streptococcus sanguis

Nine strains of Streptococcus sanguis exhibited tolerance to benzylpenicillin: the growth of each strain was susceptible to penicillin with minimal inhibitory concentrations of 0.1 μg/ml or lower, but the bacteriolytic and bactericidal effects were limited in each case. The tolerance of these bacteria was also reflected in the large discrepancies between the minimal inhibitory and minimal bactericidal concentrations for benzylpenicillin. The hypothesis that a natural deficiency of endogenous murein hydrolase (autolysin) in this species accounts for the penicillin tolerance was tested by using a heterologous murein hydrolase, the C-phage-associated lysin. In seven of the strains, addition of the lysin to the culture together with penicillin or other cell wall inhibitors resulted in lysis and rapid loss of viability. The enzyme alone did not appreciably affect normally growing cultures. The irreversible effects of penicillin plus lysin were drastically reduced in the presence of the bacteriostatic agents chloramphenicol and cerulenin. Speculations based on experiments are presented for the mechanisms by which penicillin treatment sensitizes these bacteria to an exogenous lytic enzyme. Similar phenomena requiring cooperation of host factors and penicillin may occur during infection, since somewhat similar although less pronounced results were obtained by addition of human lysozyme to penicillin-treated S. sanguis.     

Susceptibilities of oral and nasal isolates of Streptococcus mitis and Streptococcus oralis to macrolides and PCR detection of resistance genes

The susceptibility of viridans group streptococci to macrolides was determined. Thirteen isolates (17%) were resistant to erythromycin. Five strains carried an erm gene that was highly homologous to that in Tn917. Four strains had mefE genes that coded erythromycin efflux ability.     

Efficacy of Levofloxacin for Experimental Aortic-Valve Endocarditis in Rabbits Infected with Viridans Group Streptococcus or Staphylococcus aureus

Levofloxacin is among the more active fluoroquinolones against streptococci and staphylococci. It is effective against moderately severe infections caused by these organisms, but its efficacy in the treatment of bacteremia and serious infections such as endocarditis is not well defined. We compared the efficacy of levofloxacin to those of standard agents in the rabbit model of aortic-valve endocarditis caused by fluoroquinolone-susceptible strains including a penicillin-susceptible strain of Streptococcus sanguis, a penicillin-resistant strain of Streptococcus mitis, a methicillin-resistant strain of Staphylococcus aureus, and a methicillin-susceptible strain of S. aureus. Levofloxacin administered intramuscularly at dosages of 20 to 40 mg/kg of body weight twice daily (b.i.d.) was completely ineffective against the penicillin-susceptible strain, with mean vegetation titers after 3 days of therapy not statistically significantly different from those for controls. Levofloxacin was no more effective than penicillin against the penicillin-resistant strain. Levofloxacin administered for 4 days at a dosage of 20 mg/kg b.i.d. was at least as effective as vancomycin administered intravenously at a dosage of 25 mg/kg b.i. d. against the methicillin-resistant S. aureus strain and was as effective as nafcillin administered intramuscularly at 100 mg three times daily against the methicillin-susceptible strain. Emergence of resistance to levofloxacin in vitro was less likely to occur than resistance to ciprofloxacin, and resistance to levofloxacin was not observed in vivo. Levofloxacin-rifampin combinations were antagonistic in vitro and in vivo. Levofloxacin was highly effective as a single agent against experimental staphylococcal endocarditis but was surprisingly ineffective against streptococcal endocarditis, suggesting that it has a potential role as treatment for serious S. aureus but not viridans group streptococcal infections in humans.     

Streptococcus bovis Bacteraemia Requires Rigorous Exclusion of Colonic Neoplasia and Endocarditis

Twelve patients presented to the hospitals of the Auckland HospitalBoard with bacteraemia caused by Streptococcus bovis in theyears 1979–84. Ten had endocarditis, affecting homo-graftvalves in two cases and the tricuspid valve in one case. Ofnine patients who underwent investigation of the large bowel,only one did not have a colorectal tumour. Three had colonicadenocarcinoma and three had colorectal villous adenoma. Two,including a patient with acute hepatic failure from alcoholiccirrhosis, had colonic adenomata. Colonoscopy provided a tissuediagnosis of colorectal neoplasia despite negative radiologicalstudies in three patients. Bacteraemia due to S. bovis shouldprompt rigorous investigation to exclude both endocarditis andtumours of the large bo