Effect of Type-Specific Active Immunization on the Development and Progression of Experimental Pseudomonas aeruginosa Endocarditis

Rabbits with intracardiac catheters were immunized with heat-killed Pseudomonas aeruginosa or saline and challenged with either 10(9) (high inoculum) or 10(7) (low inoculum) pseudomonas. Immunization did not decrease the incidence of endocarditis when compared with controls, but it did significantly prolong survival. The longer survival of immunized rabbits after high-inoculum challenge was not due to prolongation of the course of endocarditis but to type-specific protection from early, overwhelming bacteremia. However, after low-inoculum challenge there were no early deaths and there was a significantly (P < 0.01) longer survival of immunized (17.4 days) than unimmunized (10.6 days) animals dying of endocarditis. Increased survival was associated with higher total and 2-mercaptoethanol-resistant hemagglutinating antibody titers 1 week after challenge in immunized as compared with unimmunized rabbits. Early (48 h after challenge) vegetation colonization was also significantly (P < 0.05) greater after type-specific as opposed to non-type-specific or saline immunization and low-inoculum challenge. However, whereas 67% of type-specifically immunized rabbits had colonized vegetations at 48 h, only 38.9% died with bacteremic endocarditis. Another 19.2% of immunized rabbits had vegetations colonized with > 10(5) colony-forming units of pseudomonas at elective sacrifice 2 weeks after challenge but no bacteremia; no unimmunized rabbit exhibited similar late colonization. Preexisting antibody may be important in the pathogenesis of pseudomonas endocarditis in drug addicts, and its presence may explain the subacute and often protracted course of the disease.     

Antibodies to capsular polysaccharides are not protective against experimental Staphylococcus aureus endocarditis.

The protective efficacy of antibodies to the Staphylococcus aureus capsular polysaccharide was examined in a rat model of catheter-induced endocarditis. Capsular antibodies were induced either by active immunization with killed S. aureus or by passive immunization with hyperimmune rabbit antiserum to S. aureus. Control rats were injected with phosphate-buffered saline or passively immunized with normal rabbit serum or rabbit antiserum to a nonencapsulated strain. Animals with indwelling catheters were challenged intravenously with 5 x 10(4) to 4 x 10(6) CFU of the homologous S. aureus strain (capsular serotype 5 strain Reynolds or serotype 1 strain SA1 mucoid). Both immunized and control rats developed S. aureus endocarditis. The numbers of S. aureus cells recovered from the blood and aortic valve vegetations of immunized rats were similar to those of control rats, indicating that capsule-specific antibodies were not protective. To determine whether the presence of an indwelling catheter interfered with antibody-mediated protection against S. aureus endocarditis, catheters were removed 2 h after insertion in additional groups of rats. An inoculum of 10(8) CFU of strain Reynolds was needed to provoke endocarditis in rats catheterized for 2 h, compared with 5 x 10(4) CFU for rats with indwelling catheters. Passively transferred capsular antibodies were not protective since both immunized and nonimmunized animals developed endocarditis, and quantitative cultures of blood and valvular vegetations revealed no differences between immunized and control animals. The findings of this study indicate that antibodies to the capsular polysaccharide are not protective in the rat model of experimental S. aureus endocarditis.     

Role of monocytes in experimental Staphylococcus aureus endocarditis

In the pathogenesis of bacterial endocarditis (BE), the clotting system plays a cardinal role in the formation and maintenance of the endocardial vegetations. The extrinsic pathway is involved in the activation of the coagulation pathway with tissue factor (TF) as the key protein. Staphylococcus aureus is a frequently isolated bacterium from patients with BE. We therefore investigated whether S. aureus can induce TF activity (TFA) on fibrin-adherent monocytes, used as an in vitro model of BE. We also assessed in vivo in rabbits with catheter induced vegetations, the effect of S. aureus infection on vegetational TFA. In vitro experiments showed that adherent S. aureus induced TFA on fibrin-adherent monocytes which was optimal at a bacterium/monocyte ratio of 1 to 1. Monocyte damage occurred when this ratio exceeded 4 to 1 (visually) or 6 to 1 (propidium iodide influx) Consequently, TFA decreased. In vivo S. aureus led to very high bacterial numbers in the vegetations and a significant increase of their weight. However, TFA of infected vegetations was the same as of sterile ones. This may be due to the high bacteria to monocyte ratio as well as bacterium-induced monocyte damage. Teicoplanin treatment of infected rabbits reduced bacterial numbers in the blood and in the vegetations. Two-day treatment resulted in an increase of vegetational TFA, but after four-day treatment vegetational TFA dropped, most probably due to a suboptimal bacterium/monocyte ratio. S. aureus endocarditis in etoposide (Vepesid)-treated rabbits, leading to a selective monocytopenia, caused a rapid death of the animals. In these rabbits no vegetations were found at all. We conclude that, like Streptococcus sanguis and Staphylococcus epidermidis, S. aureus is able to induce TFA in fibrin-adherent blood monocytes. In addition, monocytes have a protective effect during the course of S. aureus endocarditis.     

Protective efficacy of antibodies to the Staphylococcus aureus type 5 capsular polysaccharide in a modified model of endocarditis in rats.

The protective efficacy of antibodies to the Staphylococcus aureus type 5 capsular polysaccharide (CP5) was examined in a modified model of catheter-induced endocarditis. Rats were catheterized by surgically passing a polyethylene catheter through the right carotid artery and aortic valve into the left ventricle. The following day, the rats were injected by the intraperitoneal (i.p.) route with immunoglobulin G (IgG) purified from nonimmunized rabbits or from rabbits immunized with a conjugate vaccine composed of CP5 and CP8 linked covalently to recombinant Pseudomonas aeruginosa exotoxoid A. One day after passive immunization, the animals were challenged i.p. with one of three serotype 5 S. aureus isolates (strain Reynolds, Lowenstein, or VP) or nontypeable strain 521. Protection was evaluated by comparing quantitative cultures of blood, endocardial vegetations, and kidneys from control and immune animals. For experiments performed with S. aureus Reynolds and Lowenstein, rats given capsular antibodies (645 microg of CP5-specific IgG) showed a significantly (P < 0.05) lower prevalence of endocarditis than rats injected with nonimmune IgG. Similarly, quantitative cultures of the blood, kidneys, and aortic valve vegetations revealed that fewer S. aureus cells were recovered from rats given capsule-specific IgG than from rats administered nonimmune IgG. Rats challenged with strain VP were protected with 1.145 mg of CP5-specific IgG. Capsular antibodies did not protect against infection elicited by a nontypeable strain. These results demonstrate that capsular antibodies elicited by immunization with a polysaccharide-protein conjugate vaccine protect experimental animals against serotype 5 S. aureus infection in a modified model of endocarditis.     

Experimental Streptococcus sanguis endocarditis: immune complexes and renal involvement.

An increase of agglutinins and circulating immune complexes (CIC) was observed in the sera of rabbits after the induction of bacterial endocarditis (BE) by intravenous injection of live streptococci into animals with a non-bacterial thrombotic endocarditis. Rabbits immunized for 12 days with heat-killed streptococci developed higher levels of agglutinins and CIC than did those unimmunized during BE. The levels remained elevated in the immunized rabbits after the induction of streptococcal endocarditis. In rabbits with very high levels of CIC complement activation occurred in vivo, possibly via the classical pathway. Kidneys from both unimmunized and immunized rabbits with streptococcal endocarditis contained bacteria and also showed inflammatory reactions but only the renal tissue of immunized rabbits with endocarditis showed a little mesangial deposition of immunoglobulin. The findings in this study indicate that in rabbits, renal inflammation due to BE is initiated more often by septic emboli than by immune complex deposition, at least during the first 12 days after infection.     

Detection of anti-teichoic acid immunoglobulin G antibodies in experimental Staphylococcus epidermidis endocarditis.

An enzyme-linked immunosorbent assay (ELISA) was developed for the detection of rabbit immunoglobulin G (IgG) antibodies to purified cell wall teichoic acids from the Staphylococcus aureus Lafferty strain and three strains of coagulase-negative staphylococci. Significant immunological cross-reactivity occurred only between the teichoic acid of S. aureus and one coagulase-negative preparation. The ELISA was used to determine the serum IgG response to Staphylococcus epidermidis in a rabbit model of aortic valve endocarditis. Blood samples were drawn before inoculation and then every 5 days until death or sacrifice at 32 to 35 days postinoculation. Valve vegetations were culture positive at autopsy in 16 (59%) of the 27 catheterized rabbits. Antibody titers in this culture-positive group and the culture-negative group began to rise as early as day 6. Although both groups demonstrated an antibody response, the culture-positive group attained a significantly higher titer on days 26 and 31. Antibodies also rose in a control group of rabbits without a heart catheter but which were inoculated with bacteria. Again, the antibody titer was significantly less than that for the culture-positive group. This ELISA may be useful for the diagnosis of coagulase-negative staphylococcal infections in humans.     

Induction of experimental endocarditis by continuous low-grade bacteremia mimicking spontaneous bacteremia in humans

Transient high-grade bacteremia following invasive procedures carries a risk of infective endocarditis (IE). This is supported by experimental endocarditis. On the other hand, case-control studies showed that IE could be caused by cumulative exposure to low-grade bacteremia occurring during daily activities. However, no experimental demonstration of this latter possibility exists. This study investigated the infectivity in animals of continuous low-grade bacteremia compared to that of brief high-grade bacteremia. Rats with aortic vegetations were inoculated with Streptococcus intermedius, Streptococcus gordonii or Staphylococcus aureus (strains Newman and P8). Animals were challenged with 10(3) to 10(6) CFU. Identical bacterial numbers were given by bolus (1 ml in 1 min) or continuous infusion (0.0017 ml/min over 10 h). Bacteremia was 50 to 1,000 times greater after bolus than during continuous inoculation. Streptococcal bolus inoculation of 10(5) CFU infected 63 to 100% vegetations compared to 30 to 71% infection after continuous infusion (P > 0.05). When increasing the inoculum to 10(6) CFU, bolus inoculation infected 100% vegetations and continuous infusion 70 to 100% (P > 0.05). S. aureus bolus injection of 10(3) CFU infected 46 to 57% valves. This was similar to the 53 to 57% infection rates produced by continuous infusion (P > 0.05). Inoculation of 10(4) CFU of S. aureus infected 80 to 100% vegetations after bolus and 60 to 75% after continuous infusion (P > 0.05). These results show that high-level bacteremia is not required to induce experimental endocarditis and support the hypothesis that cumulative exposure to low-grade bacteremia represents a genuine risk of IE in humans.     

Natural history of aortic valve endocarditis in rats.

Sterile aortic vegetations were produced in rats by introducing a polyethylene catheter through the right carotid artery. The catheter was either left in place throughout the experiments or removed before bacterial challenge. Bacterial endocarditis was uniformly produced by intravenous injection of 10(7) colony-forming units of Staphylococcus aureus or Streptococcus intermedius, whether the catheter was left in place or removed. However, in rats with the catheter left in place, bacterial multiplication within the vegetations with both strains was accelerated, and mortality from Staphylococcus aureus infection was increased. Using 10(7) colony-forming units of serum-resistant Escherichia coli as a test microorganism, we found a marked difference in the production of endocarditis depending upon whether the catheter was left in place or removed before injection; only those animals infected with the catheter in place developed infection. From these experiments in rats, it was evident that the presence of a foreign body has a considerable influence on the ability of bacteria to grow within an intravascular vegetation. In addition, a striking difference in the virulence of the three strains studied was established; Staphylococcus aureus was the most, and E. coli the least, pathogenic.     

Comparative opsonic and protective activities of Staphylococcus aureus conjugate vaccines containing native or deacetylated Staphylococcal Poly-N-acetyl-beta-(1-6)-glucosamine

Staphylococcus aureus and Staphylococcus epidermidis both synthesize the surface polysaccharide poly-N-acetyl-beta-(1-6)-glucosamine (PNAG), which is produced in vitro with a high level (>90%) of the amino groups substituted by acetate. Here, we examined the role of the acetate substituents of PNAG in generating opsonic and protective antibodies. PNAG and a deacetylated form of the antigen (dPNAG; 15% acetylation) were conjugated to the carrier protein diphtheria toxoid (DT) and used to immunize animals. Mice responded in a dose-dependent fashion to both conjugate vaccines, with maximum antibody titers observed at the highest dose and 4 weeks after the last of three weekly immunizations. PNAG-DT and dPNAG-DT vaccines were also very immunogenic in rabbits. Antibodies raised to the conjugate vaccines in rabbits mediated the opsonic killing of various staphylococcal strains, but the specificity of the opsonic killing was primarily to dPNAG, as this antigen inhibited the killing of S. aureus strains by both PNAG- and dPNAG-specific antibodies. Passive immunization of mice with anti-dPNAG-DT rabbit sera showed significant levels of clearance of S. aureus from the blood (54 to 91%) compared to control mice immunized with normal rabbit sera, whereas PNAG-specific antibodies were ineffective at clearing S. aureus. Passive immunization of mice with a goat antiserum raised to the dPNAG-DT vaccine protected against a lethal dose of three different S. aureus strains. Overall, these data show that immunization of animals with a conjugate vaccine of dPNAG elicit antibodies that mediated opsonic killing and protected against S. aureus infection, including capsular polysaccharide types 5 and 8 and an untypable strain.     

Enzyme-linked immunosorbent assay for detection of Staphylococcus epidermidis antibody in experimental S. epidermidis endocarditis.

The immune response against Staphylococcus epidermidis, as determined by an enzyme-linked immunosorbent assay, was evaluated in experimental S. epidermidis infections in rabbits. Antigens from 8 of 10 clinical S. epidermidis strains detected significant antibody production in five rabbits immunized with different strains of S. epidermidis and in five of six rabbits with experimental endocarditis caused by four different strains. The antigens from two strains detected antibody production in all rabbits, and strain ATCC 14990 discriminated best between positive and negative samples. Consecutive blood samples from rabbits with endocarditis and control rabbits with bacteremia, which were successfully prevented from developing endocarditis by using prophylactic antibiotics, were examined by using an enzyme-linked immunosorbent assay and an ultrasonic extract of strain ATCC 14990 as the antigen. This assay discriminated between rabbits with endocarditis and rabbits with uncomplicated bacteremia. Antibody production was detected as early as 3 days after the onset of infection in rabbits with endocarditis.     

Anti-Gamma Globulins and Chronic Infection: Comparative Studies of the Immune Response to Various Bacteria and Gamma Globulin Preparations

A study of the relationship of clinical states associated with prolonged infection (bacterial endocarditis and osteomyelitis) and generation of serum anti-gamma globulins was made with particular reference to quantitative amounts of staphylococcal protein A in various infecting strains. No correlation between individual strain amounts of protein A and presence of anti-gamma globulins was detected. Thirty-eight rabbits were immunized intravenously with various strains of bacteria (Staphylococcus aureus, enterococci, Streptomyces viridans, pneumococci, pseudomonas, and Escherichia coli) for periods of 6 weeks, and antibacterial as well as anti-gamma globulin antibodies were assayed. No single group or strain of bacteria stood out as being more prone to produce anti-gamma globulins than others tested. Most rabbits developed anti-gamma globulins reacting with human gamma globulins, whereas the specificity for rabbit gamma globulin appeared more restricted. In 16 rabbits immunized with eight different strains of S. aureus, quantitative elevation of serum gamma globulin above 2.5 g per 100 ml often seemed to be correlated with presence of detectable serum anti-gamma globulins. By contrast 15 rabbits immunized with autologous or isologous rabbit gamma globulins in many instances developed extremely high titers of anti-gamma globulins showing primary specificity for human rather than rabbit gamma globulin. These studies further amplify the remarkably heterogeneous anti-gamma globulin reactivity associated with various types of immune response.     

Klebsiella pneumoniae type 3 fimbria-mediated immunity to infection in the murine model of respiratory disease

Type 3 fimbriae are expressed by most strains of Klebsiella pneumoniae and facilitate adherence to the basement membrane of human respiratory tissues. The ability of these appendages to stimulate a protective immune response in vivo has not been investigated. A murine model of acute pneumonia was used to determine whether the production of type 3 fimbria-specific antibodies correlated with protection against infection by K. pneumoniae. Purified fimbriae from several strains were used to immunize mice prior to challenge with a virulent strain. The immunized mice produced high titers of specific antibody and this was associated with protection against challenge with a low dose of bacteria that was lethal in unimmunized animals. However, challenge with a high number of bacteria resulted in no protection against infection.     

Effect of immunization on susceptibility to experimental Streptococcus mutans and Streptococcus sanguis endocarditis.

It has been asserted that humoral immunity is an important potentiating factor in pathogenesis of infective endocarditis, in that prior immunization to certain bacteria may predispose the host to endocarditis caused by those organisms. If so, possible future vaccination of humans with streptococcal antigens for the prevention of dental caries might increase the susceptibility of the population to streptococcal endocarditis. To examine this hypothesis further, we immunized rabbits with killed Streptococcus sanguis or Streptococcus mutans. After complement-fixing antibody had developed, the rabbits were tested for susceptibility to experimental infective endocarditis. Rabbits with high titers of complement-fixing antibody to the infecting organism developed streptococcal endocarditis less often (13%) than animals with lower titers (69%; P less than 0.0002). These findings do not support the hypothesis that pre-immunization predisposes to infective endocarditis and lend no credence to the concept that vaccination of human subjects against dental caries might increase their susceptibility to streptococcal endocarditis. On the contrary, the results of these experiments indicate that specific antibody can confer relative immunity to infective endocarditis.     

Human monoclonal antibody AVP-21D9 to protective antigen reduces dissemination of the Bacillus anthracis Ames strain from the lungs in a rabbit model

Dutch-belted and New Zealand White rabbits were passively immunized with AVP-21D9, a human monoclonal antibody to protective antigen (PA), at the time of Bacillus anthracis spore challenge using either nasal instillation or aerosol challenge techniques. AVP-21D9 (10 mg/kg) completely protected both rabbit strains against lethal infection with Bacillus anthracis Ames spores, regardless of the inoculation method. Further, all but one of the passively immunized animals (23/24) were completely resistant to rechallenge with spores by either respiratory challenge method at 5 weeks after primary challenge. Analysis of the sera at 5 weeks after primary challenge showed that residual human anti-PA levels decreased by 85 to 95%, but low titers of rabbit-specific anti-PA titers were also measured. Both sources of anti-PA could have contributed to protection from rechallenge. In a subsequent study, bacteriological and histopathology analyses revealed that B. anthracis disseminated to the bloodstream in some na?ve animals as early as 24 h postchallenge and increased in frequency with time. AVP-21D9 significantly reduced the dissemination of the bacteria to the bloodstream and to various organs following infection. Examination of tissue sections from infected control animals, stained with hematoxylin-eosin and the Gram stain, showed edema and/or hemorrhage in the lungs and the presence of bacteria in mediastinal lymph nodes, with necrosis and inflammation. Tissue sections from infected rabbits dosed with AVP-21D9 appeared comparable to corresponding tissues from uninfected animals despite lethal challenge with B. anthracis Ames spores. Concomitant treatment with AVP-21D9 at the time of challenge conferred complete protection in the rabbit inhalation anthrax model. Early treatment increased the efficacy progressively and in a dose-dependent manner. Thus, AVP-21D9 could offer an adjunct or alternative clinical treatment regimen against inhalation anthrax.     

Effect of immunization on the genesis of pneumococcal endocarditis in rabbits.

The effect of immunization with whole organisms on the development of Streptococcus pneumoniae endocarditis was examined by in vivo and in vitro methods. Immunization protected rabbits from pneumococcal endocarditis when the in vivo catheterization model was used. The inoculum size that caused endocarditis in 50% of the unimmunized rabbits was 1.1 X 10(5) colony-forming units, whereas 1.2 X 10(7) colony-forming units were required for infecting 50% of the immunized rabbits (P less than 0.001). Investigations were carried out to determine the mechanism which enabled immunization to prevent the development of pneumococcal endocarditis; they indicated that a reduction in bacterial adherence could not explain this phenomenon. In vitro studies showed that subagglutinating quantities of antibody increased the adherence (P less than 0.05) of pneumococci to rabbit aortic valve cusps. The adherence ratio of pneumococci to fibrin-platelet clots was at least doubled by the presence of subagglutinating dilutions of immune sera (P less than 0.001). Further studies showed that immunoglobulin G in the immune sera accounted for this increased in vitro adherence. However, further in vivo studies showed that immunized rabbits were able to clear live pneumococci from their bloodstreams within 4.5 h, whereas unimmunized rabbits failed to clear the organism within 24 h.     

Immunization with the N-terminal portion of Treponema pallidum repeat protein K attenuates syphilitic lesion development in the rabbit model

When used as an immunogen, Treponema pallidum repeat protein K (TprK) has been shown to attenuate syphilitic lesions upon homologous intradermal challenge in the rabbit model. To further explore this protein as a potential vaccine component, we sought to identify the immunogenic regions of TprK. The abilities of three recombinant peptides encompassing TprK to elicit T- and B-cell responses and to protect against challenge were examined. All three fragments elicited proliferative responses from splenocytes taken from infected rabbits. However, enzyme-linked immunosorbent assays indicated that only fragments 1 and 3 were consistently recognized by antisera from infected rabbits. Each fragment was also used to immunize rabbits that were subsequently challenged intradermally with infectious T. pallidum. All lesions on unimmunized control rabbits ulcerated and contained treponemes, while the lesions on rabbits immunized with fragment 1 were the least likely to have detectable treponemes (25%) and the least likely to ulcerate (37.5%). The lesions on rabbits immunized with fragment 3 showed intermediate results, and rabbits immunized with fragment 2 were the most likely of all those on immunized rabbits to have detectable treponemes (91.7%) and to ulcerate (66.7%). These results demonstrate that epitopes in fragment 1 are recognized by T cells and antibodies during infection and that immunization with this portion of TprK most effectively attenuates syphilitic lesion development.     

Role of the Vegetation in Experimental Streptococcus viridans Endocarditis

This study examines the role of the vegetation in catheter-induced experimental endocarditis in predisposing to bacterial colonization of cardiac valves and in influencing the course of the disease and response to penicillin therapy. Platelet-fibrin vegetations developed at areas of valvular trauma and were colonized when Streptococcus viridans were injected intravenously. Pretreatment with warfarin prevented vegetation formation, but animals still developed endocarditis at the same rate after injection of 10(6)S. viridans. The course of the disease in anticoagulated animals was more explosive, as determined by a more rapid rise in fever and level of bacteremia. Mean survival was shorter in anticoagulated rabbits (7 versus 12.7 days). Large vegetations containing 10(9)S. viridans/g were found in control animals, whereas anticoagulated rabbits developed only microscopic deposits. Large vegetations required a longer duration of penicillin therapy to sterilize than the infected valves of the anticoagulated group (7 versus 3 days). Therefore, a preformed platelet-fibrin deposit is not a prerequisite for bacterial colonization of cardiac valves. After infection, the vegetation is an important factor in determining the subacute course of disease and resistance to penicillin therapy.     

Immunization with Treponema pallidum endoflagella alters the course of experimental rabbit syphilis.

Rabbits were immunized over a 32-week period with a total of 450 micrograms of purified Treponema pallidum endoflagella. As measured by enzyme-linked immunosorbent assay, sera from immunized rabbits had antiendoflagellar antibody titers that were fivefold greater than titers of sera from infected immune rabbits and patients with secondary disease. Sera from all immunized animals possessed complement-dependent treponemicidal activity as measured by in vitro immobilization. Immunized animals challenged with virulent T. pallidum were not protected from symptomatic infection but showed an altered course of lesion development.     

Role of granulocytes and monocytes in experimental Staphylococcus epidermidis endocarditis.

The role of granulocytes and monocytes during the induction and course of Staphylococcus epidermidis endocarditis was investigated by the selective depletion of monocytes with the drug VP16-213 and of both granulocytes and monocytes with nitrogen mustard. The induction of endocarditis was influenced only by the depletion of monocytes: the 50% infective dose differed significantly, being 3.4 X 10(5) CFU in control rabbits and 3.4 X 10(4) CFU in the monocyte-depleted rabbits, whereas no significant differences were found between the latter and those depleted of both granulocytes and monocytes. Also, control rabbits injected with 10(6) or 10(7) CFU had a significantly higher incidence of sterile vegetations than did rabbits selectively depleted of granulocytes or monocytes. Compared with baseline values, mean monocyte numbers at the time of bacterial inoculation were significantly increased in control rabbits whose vegetations remained sterile, whereas this effect was not seen in rabbits whose vegetations became infected. The course of the endocarditis appeared to be significantly influenced by both granulocytes and monocytes. Comparison showed that a decrease of the same numbers of these cells per microliter of blood was accompanied for the monocytes by an approximately fourfold higher increase of the number of staphylococci in the vegetations. The correlation between the number of granulocytes and of monocytes on the one hand and the number of staphylococci in the vegetations on the other was not substantially influenced by the duration of the disease or the number of staphylococci injected to induce the endocarditis. The number injected proved to be significantly correlated with the number of staphylococci in the vegetations. In rabbits with numbers of CFU per gram of vegetation exceeding 10(7), blood cultures were usually positive. This finding applied rarely to control rabbits, but generally to drug-treated rabbits. In the latter animals a significant correlation between the number of staphylococci in the vegetations and in the circulation was found. We conclude that only monocytes have a measurable effect on the induction of Staphylococcus epidermidis endocarditis but during its course both granulocytes and monocytes keep the endocardial infection in check.     

Enhancement of experimental bacteremia and endocarditis caused by dysgonic fermenter (DF-2) bacterium after treatment with methylprednisolone and after splenectomy.

The dysgonic fermenter-2 bacterium is a newly recognized fastidious gram-negative bacillus that causes bacteremia and sometimes endocarditis in immunocompromised persons after they are bitten by dogs. To develop an experimental model of this infection, we placed polyethylene catheters across the aortic valves of New Zealand white rabbits, which were inoculated intravenously the next day with dysgonic fermenter-2 bacteria. After 1 week, the rabbits were killed and the endocardial vegetations were homogenized for quantitative culture. Large inocula (1.3 X 10(10) to 2.1 X 10(10) viable bacteria) were required to produce infected vegetations. All infected rabbits had negative blood cultures at the time of autopsy and most developed serum agglutinins against dysgonic fermenter-2 bacteria. Three daily injections of methylprednisolone (30 mg/kg), starting the day before inoculation, significantly increased the incidence of endocarditis and the number of bacteria per gram of infected vegetation (P less than 0.05). Treatment with methylprednisolone prolonged the initial bacteremia and caused significant increases in the numbers of bacteria per gram of blood, spleen, and liver compared with those of untreated controls (P less than 0.05). Rabbits that had previously undergone splenectomy showed prolongation of the initial bacteremia but no significant increase in the incidence of infected vegetations. These results showed that the dysgonic fermenter-2 bacterium is a pathogen that causes endocarditis in rabbits but that it requires a large inoculum and produces blood culture-negative infections. Treatment with methylprednisolone enhances infection by prolonging the initial bacteremia and probably by diminishing bactericidal activity in the vegetations.