Heptavalent pseudomonas vaccination in seriously burned children

Heptavalent Pseudomonas Vaccination in Seriously Burned Children By JAMES A. O'NEILL, JR., F. CARTER NANCE, AND MYRONW. FISHER

I

N 1961 THE MAIN CAUSE of death from thermal injury was identified as invasive burn wound sepsis due to Pseudomonas aeruginosa.’ Shortly after the definition of this problem, new approaches to infection control were investigated and the concept of topical therapy emerged. In particular, topical use of Mafenide cream resulted in considerable improvement in survival, which was especially noteworthy in childhood.2 Other topical agents, such as 0.5% silver nitrate, Gentamycin, and silver sulfadiazine, have been introduced, but the improvement in survival seen with all effective agents has not been impressive in individuals with burn injuries exceeding 60% of body surface or in any patient with established burn-wound sepsis. Although seen less commonly than before, invasive Pseudomonas infection is still a significant problem. Several studies have shown that immunoglobulins fall during the early postburn period, suggesting that an immunologic approach to infection control might be another reasonable approach to the problem.3‘s Alexander and Moncrief demonstrated some impairment of the ability to produce antibodies to a primary antigenic stimulus in experimental animals with large burns and especially when the antigenic challenge was given a few days following burning rather than immediately.8 Kefalides noted that survival was improved in children who received pooled-plasma-source gamma globulin.7 In 1968, Feller reported the combined use of a monovalent Pseudomonas vaccine and hyperimmune plasma and indicated that the number of septicemias diminished markedly.8 However, the monovalent vaccine has limited immunologic coverage. In 1969, Fisher et al. at the Parke-Davis Research Laboratories reported on the development of an immunotype schema for Pseudomonas based on challenge protection in mice. a Seven serotypes were found to cross-react with over 95% of all known pathogenic strains of Pseudomonas. A heptavalent vaccine was then constructed as purified soluble antigens derived from killed bacteria constituting the seven serotypes described by Fisher. Alexander had previously suggested the feasibility of this approach. lo Extensive studies in animals and normal volunteers provided information indicating that the vaccine could be given safely and that it could evoke satisfactory antibody response. The present study details preliminary experience with this heptavalent PseuFrom Department of Surgery, Louisiana State University Medical School, New Orleans, and Medical and Scientific Affairs Division, Parke-Davis and Company, Detroit, Mich. Presented before the American Pediatric Surgical Association, Hamilton, Bermuda, April 22-24, 1971. JAMES A. O’NEILL, JR., M.D.: Professor of Surgery, Surgeon-in-Chief, Vanderbilt University Children’s Medical Center, Nashville, Tenn. F. CARTER NANCE, M.D.: Associate Professor of Surgery, Louisiana State University Medical Center, New Orleans, La. MYRON W. FISHER, PH.D.: Director, Bacteriology and Mycology Research, )Pnrke-Davis and Company, Detroit, Mich.

JOURNALOF PEDIATRICSURGERY,VOL. 6, No. 5 (OCTOBER). 1971

547

548

O’NEILL, NANCE, AND FISHER

domonus vaccine in a series of burned children in terms of its safety and its ability to produce antibodies, and, in a limited way, its ability to prevent invasive infection. MATERIALS AND METHODS Thirty-two consecutive children with thermal injuries exceeding 10% of body surface were included in the study. All patients received topical chemotherapy either with Mafenide or silver sulfadiazine cream. Most also received penicillin initially and several individuals received other antibiotics during their course on specific culture indication. On admission to the hospital, all patients were coded in a double-blind, randomized fashion and were then given either saline placebo or heptavalent Pseudomonas vaccine supplied by Parke-Davis. The first two patients received a dose of 10 Kg/kg of body weight but all subsequent patients have received 25 pg/kg. The vaccine was given on admission to the hospital and then every 4 days for two doses and subsequently once a week for a maximum total of five doses. Depending upon the time of healing or skin coverage, patients received from two to five doses. All individuals were carefully observed for 48 hr after each injection for any evidence of local or systemic reactions. Blood samples were drawn for antibody titer prior to the initial injection and at weekly intervals thereafter. Immunoglobulin M ( IgM) and immunoglobulin G( IgG) titers for each of the seven major serotypes of Pseudomonas were measured on each serum sample by a passive hamagglutination technique by Dr. H. B. Devlin at the Parke-Davis Research Laboratories. Wound, blood, and other indicated cultures were performed at frequent intervals and all Pseudomonas isolates were typed. Survival, cause of death, and incidence and type of infection were noted. All data were subjected to random block analysis of variance.

RESULTS Of the 32 patients included in the study, 19 received placebo and 13 received vaccine. Age range in the entire group was from 5 mo to 10 yr. Average age in the controls was 4.6 yr and 4.8 yr in those vaccinated. Burn injuries in the entire group ranged from 12 to 75% of body surface. The mean total burn in the control group was 23% and 24% in the vaccinated patients, A proportional distribution of Mafenide- and silver-sulfadiazine-treated patients were in each group. One patient who received saline placebo developed mild redness around the injection site, which lasted about 2 hr, and he also had a two-degree elevation in temperature during that morning. Five of the 13 vaccine-treated patients developed similar mild local reactions and a transient elevation in temperature. However, the latter factor is difficult to evaluate in burned individuals since they tend to be febrile much of the time. Saline placebo did not produce any elevation of antibody titers in the control patient group, as would be expected. An occasional patient in this group showed evidence of self-immunization with a response to a particular Pseudomonas serotype, which was usually present on culture. The patients who received vaccine responded vigorously, and the response was usually evident within 4 days of the initial injection, although usually not before that time ( Fig. 1) . The vaccinated group had a highly statistically significant rise in antibody titers for all seven serotypes in response to immunization when compared with the controls. (Fig. 2). There were some exceptions to this, however. The first two patients received a vaccine dose of 10 pg/kg and, although their antibody titers were probably protective, the response was lower

VACCINATION

IN BURNED

549

CHILDREN

PT. No. 89 Age-S Total Burn 407x,, 2S% 3rd” Pseud 1 type 81922948 1024 512 256 128 64 32 16 8 4 4

00

)II---

1 4 7 10 14 fW4 4 DAYSPOSTBURN

7

24

Fig. 1 .-IgG response for all seven serotvpes in response to 25 &kg of heptavnlent Pseudomonas antigen administered on the first postburn day. Note that it takes G-6 davs for active immunization to occur but that response is excellent bv this time. When infection does occur, the tvpe-specific IgG response is accentuated.

and less prolonged than for the remaining patients who received 25 kg/k%. Also, patients who were immunized later than the fourth postburn day did not respond in uniform fashion. Although the response was good in one patient whose immunization began on the 17th postburn day, three others who began immunization on the fifth, ninth, and tenth days did not respond well. Furthermore, a statistical comparison of control and vaccinated patients injected later than the fourth postburn day showed no significant difference between the two groups’ subsequent antibody titers. Those patients who received 25 &g/kg of vaccine hy the fourth postburn day uniformly responded with substantial elevations in antibody titers. The highest level achieved was for type 1 followed in order by types 2,4,5, 3,6, and 7. Wound cultures grew out Pseudomonas organisms in four of the I.9 control patients and seven of the 13 patients who received vaccine. Serotypes 1, 2, 3, 6 and 7 were encountered, but types 2 and 3 were the most frequent ones to appear in these patients. Those patients who had known Pseudomorzas colonization tended to have a more marked rise in antibody titer in response to \.accinr: administration ( Fig. 1) , COMMENTS

Since invasive Pseudomonas burn-wound sepsis continues to be a problem in large burn injuries despite the use of topical agents, it was decided to investigate the possible contribution of immunotherapy. Previous experimental studies

550

Fig. 2.-Graph outlining the difference between the highest postimmunization titer and the preimmunization titer for all 32 patients for the seven serotypes. Note the significantly higher levels of antibody response in the vaccinated patient group.

O’NEILL,

TYPa

NANCE, AND FISHER

1 WEAN CNANQE IN TITER IN RESPONIE

TO VACCINE

by Millican and Rust, among others, have shown that immunization will provide protection against a challenge of homologous strain of P.seudomonas.11 Nonrandomized clinical studies by Alexander have also suggested that immunization may provide effective protection in burned patients, who tend to have a lessened antibody response as a result of their injury. These studies, as well as those of Alexander, indicate that the use of purified Pseudomonas antigens is safe. 12 Approximately one half of the patients who received 25 pg/kg of vaccine intramuscularly developed mild, transient, local erythematous, and systemic febrile reactions. No reaction was severe enough to require any sort of treatment. Even though some patients who had Pseudomonas cultured from their wounds demonstrated elevated antibody titers as evidence of self-immunization, this response was limited and inconstant. The levels of antibody prior to immunization best indicated this phenomenon. For example, one patient with a 20% burn was admitted on the seventh postburn day and had a preimmunization type 2 IgG level of 1:256, and Pseudomonas type 2 was cultured from her wound. However, any patient’s ability to react to his own infecting antigen would be expected to diminish as the size of the injury increased. There was no difference in the degree of antibody response as related to age within the limits of this study. However, no infants younger than 5 mo were studied. Dosage did appear to make a difference. It appears that 25 @g/kg of antigen will regularly provoke a good response but lesser doses will not. This is twice the dose required to provoke an adequate response in normal subjects. Although Alexander’s studies indicate that the vaccine may not be as effective if given after the sixth postburn day, the present studies in children indicate that antibody response may be limited if vaccine is not administered by the fourth postburn day. l2 Antibody levels diminish rapidly, most likely on the basis of loss of gamma globulin into the burn wound. It appears that weekly injections are needed in order to maintain protective levels of antibody. An IgG antibody titer of 1.8 or higher is probably protective according to the studies of Alexander.12

551

VACCINATIONIN BURNEDCHILDREN

The matter of protective levels of antibody must be qualified from two points of view. First, even though both IgM and IgG antibodies may support bacterial killing by neutrophils, IgG probably plays a greater role since it is more diffusable in tissue. The present studies included both IgM and IgG determinations initially, and the responses were essentially parallel although the actual degree of response was, of course, of greater magnitude with respect to IgM. Lately, only IgG titers have been studied for this reason. Second, even though a patient may have achieved high levels of antibody production, this may not always be sufficient since the humoral response is only one, albeit important, portion of the overall immunologic chain of events involved in bacterial killing. There is good evidence that some patients with large burns have additional defects in neutrophil function such that they are incapable of engulfing and/or killing bacteria.‘3 Vaccine failure then may result from inadequate dosage, late administration that results in poor antibody response, administration to a patient who already has established sepsis, or associated impairment in neutrophil function even in the presence of adequate levels of antibody. It seems quite clear that, regardless of the classification utilized, invasive infection from Pseztdomonas is type-related. Although some cross-reactivity certainly occurs, many immunotypes are distinctly different. On this basis, it seems most reasonable to use a polyvalent vaccine that is known to contain virtually all known pathogenic serotypes for stimulation of active immunity. The good results obtained with Feller’s monovalent vaccine may be more related to his use of hyperimmune plasma in association with the vaccine.* The use of agents for passive immunization, especially in patients with established infection, or those seen several days following injury, probably has real merit. It is also quite possible that the present heptavalent vaccine might require alteration to include newly discovered pathogenic serotypes. It is not possible to tell from these preliminary studies how effectively the vaccine prevented infection. The aim was to study safety and antibody response primarily. More patients with large burn injuries must be studied to determine the vaccine’s effect on mortality. It is encouraging to note that two patients in this series with large burn injuries, extensive Pseudomotlas colonization, and Pseudomonas septicemia survived. The vaccine would not be expected to affect the rate of colonization but it would be hoped that: if given early enough, it would affect the rate of sepsis and the response to sepsis if it occurs, presuming that other aspects of the immune response are operative. Early results are encouraging and warrant further investigation and use of this vaccine, especially in patients with large burn injuries. SUMK~ARY Thirty-two infants and children with burn injury were given either saline placebo or a heptavalent Pseudomonas vaccine in a double-blind, randomized fashion. Thus far the vaccine has proved to be safe and effective in terms of invoking a significant rise in antibody titers for all of the seven major serotypes, although greater for some than for others. The vaccine must be given in ade-

552

O'NEILL,NANCE,AND

FISHER

quate dosage and, preferably, prior to the fifth postburn day in children. There appears to be a relationship between the Pseudomonas serotype, antibody titer, and resistance to infection, but more patients with large burn injuries must be studied to prove this conclusively. Preliminary resuIts are encouraging. REFERENCES 1. Teplitz, C., and Lindberg, R. B.: Burn wound sepsis: a postmortem evaluation of the burn death. U.S. Army Surg. Res. Unit Annual Report, FY 1962, BAMC, Fort Sam Houston, Tex. 2. Pruitt, B. A., Jr., O’Neill, J. A., JT., Moncrief, J. A., and Lindberg, R. B.: Successful control of burn-wound sepsis. JAMA 203: 1054, 1968. 3. Arturson, C., Johansson, S. G. O., Hogman, C. F., and Killander, J.: Changes in immunoglobulin levels in severely burned patients. Lancet 1:546, 1969. 4. Fox, J. E., and Lowbury, E. J.: Immunity and antibody to Pseudomonas pyocyonen in rabbits. J. Path. Bact. 65:533, 1953. 5. Munster, A. M., Hoagland, H. C., and Pruitt, B. A., Jr.: The effect of thermal injury on serum immunoglobulins. Ann. Surg. 172:965, 1970. 6. Alexander, J. W., and Moncrief, J. A.: Alterations of the immune response following severe thermal injury. Arch. Surg. 93:75, 1966. 7. Kefalides, N. A., Arana, J. A., Bazan, A., Velarde, N., and Rosenthal, S.: Evaluation of antibiotic prophylaxis and gamma

globulin, plasma, albumin and saline solution therapy in severe burns. Ann. Surg. 159:496, 1964. 8. Feller, I., and Pierson, C.: Pseudomonas vaccine and hyperimmune plasma for burned patients. Arch. Surg. 97:225, 1968. 9. Fisher, M. W., Devlin, H. B, and Gnabasik, F. J.: New immunotype schema for Pseudomonas aeruginosa based on protective antigens. J. Bact. 98:8X$ 1969. 10. Alexander, J. W., Brown, W., Walker, H., Mason, A. D., Jr., and Moncrief, J. A.: Studies on the isolation of an infectionprotective antigen from Pseudomonas aeruginosa. Surg. Gynec. Obstet. 123:965, 1966. 11. Millican, R. C., and Rust, J. D.: Efficacy of rabbit Pseudomonas antiserum in experimental Pseudomonas aeruginosa infection. J. Infect. Dis. 107:389, 1960. 12. Alexander, J. W., Fisher, M. W., and MacMillan, B. G.: Immunological control of Pseudomonas infection in burn patients: a clinical evaluation. Arch. Surg. 102:31, 1971. 13. -, and Wixson, D.: Neutrophil dysfunction and sepsis in burn injury. Surg. Gynec. Obstet. 130~431, 1970.

Discussion Dr. R. E. Cross burned children?

(Boston):

What

are your indications

for using Pseudomonas

vaccine

in

Dr. OWeill: We give the Pseudomonas vaccine to children who have over 15% total body burns, in whom we feel there is a chance of mortality. We are limiting use of the vaccine to this group because the vaccine still is experimental. Dr. T. Boles (Columbus): I think this is a very important bit of work, and congratulate Dr. O’Neill on presenting it, The problems in the management of bum sepsis are certainly far from over. Antibiotics have been quite disappointing, as all of you know. Even the topical agents ( silver nitrate, sulfamylon, silver sulfadiazine, etc. ), although dramatically helping the smaller bums particularly, have not obviated mortality in the really extensive bums. Actually, the data thus far compiled by the National Bum Information Exchange indicate

VACCINATION

that mortality

IN BURNED

CHILDREN

figures in burned

children

553 have not been affected

by any type of local therapy.

Now whether or not this will ultimately prove to be true is difficult to say, but at the moment this is the way the figures read. Therefore, this type of approach to enhance the body’s defenses certainly has much to recommend it. I would like to ask Dr. O’Neill whether or not he has tried or feels there may be a place for passive immunization, until the active immunity has a chance to build up.

Dr. OWeill: We did not discuss passive immunization in this presentation. As Dr. Cross said, you have to save a little bit for the next meeting. We feel that there is without question a place for passive immunization. We have been using a heptavalent-fortified globulin preparation, which is prepared by giving this antigen to volunteers. Certainly for the patient who is in jeopardy or the person who comes in late, passive immunization is a very valuable adjunct, so I am glad you brought up the question. Dr. L. Lither (Phoenix): I also want to commend Dr. O’Neill for this unique method of overcoming a significant problem, Pseudomonas sepsis, in burn therapy. There will always be a quest for diminishing the Pseudomonas overgrowth by the use of topical agents. I would like to call your attention to material we have used over the past few years. We have been impressed with povidone-iodine, commonly known to you as Betadine, for 10 yr. Detailed data accumulated by us will be published later, but I wish to give you a preliminary report. The most severe burns were treated with a single topical agent, Betadine solution. The mortality was zero in the Betadine-treated patients and four out of 165 in the control group, which included various other topical antiseptic treatments. There was a more than 60% diminution of Pseudomonas growth in the Betadine group as compared with the controls, even though bum severity for the Betadine-treated group was significantly greater than for the control group. We hope that other burn units might undertake similar studies to enlarge the series rapidly. We are encouraged by the ease with which the Betadine solution is applied, its lack of toxicity, and the diminishing incidence of burn sepsis, specifically Pseudomonas. Dr. OWeill: I’ll make one comment about Betadine therapy of burns. Those of us who have been involved in the evaluation of topical agents have used primarily the burned-rat sepsis model. Betadine, as well as other types of iodine preparations, have proved not to be effective in the rat-burn model. In the human evaluation of such agents we must be extremely careful to categorize burns in terms of size, other complications, and the like. I am not commenting on the value of Betadine in Dr. Linkner’s series but rather cautioning you to evaluate it on the basis of a fairly reliable model in experimental animals.