Simultaneous Infection with Legionella pneumophila and Pittsburgh Pneumonia Agent Clinical Features and Epidemiologic Implications
ROBERT R. MUDER, M.D. VICTOR L. YU, M.D. RICHARD M. VICKERS, B.S. JOHN RI%,
JOHN SHONNARD, M.D. Pittsburgh, Pennsylvania
Nosocomial pneumonia caused simultaneously by two organtsms, Legionella pneumophila and the Plttsburgh pneumonia agent, was documented in seven patlents in one lnstltutton. In all seven cases, both organisms were demonstrated by lsolatlon from culture or visualization by direct Immunofluorescence. Four patients died as a result of pneumonia, including two who received erythromycln therapy. The hospital water distribution system appeared to be the reservoir for both L. pneumophlla and Plttsburgh pneumonia agent. These seven cases constituted 26.9 percent and 17.9 percent of the cases of Pittsburgh pneumonia agent and Legionnaires’ disease, respectively, at one instftutlon. Given this relatively high kcidence of dual infection, it Is likely that the mode of transmission for both organisms is identical. Dual infection may account for some cases of antibody respcnse to more than one Leglonella specks. Historical parallels of the discovery of L. pneumophila and Plttsburgh pneumonia agent are reviewed. The discovery
bacterium) as a cause of pneumonia [I] was soon followed by isolation of other organisms wlth the potential of causing human infection
From the Dapartrnantsof Medicine and Pathology, Veterans Administration Medical Center, and the University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania. Requests for reprints should ba addressedto Dr. Victor L. Yu, Infectious Disease section, Veterans Administration Medical Center, University Drive C, Pittsburgh, Pennsylvania 15240. Manuscript accepted August 31, 1982.
[2-41. Pittsburgh pneumonia agent (Legionella micdadei , Legionella pittsburghensis , Tatlockia micdadei ) shares morphologic, metabolic, and antigenic characteristics with L. pneumophila. Pittsburgh pneumonia agent, like L. pneumophila, is a cause of nosocomial pneumonia, particularly in immunocompromised patients [8,9]. Furthermore, Pittsburgh pneumonia agent has been isolated from a hospital water system known to be a reservoir for L. pneumophila [lo]. Pittsburgh pneumonia agent and L. pneumophila, therefore, appear to share epidemiologic features as well. We herein describe seven patients with simultaneous infection by L. pneumophila and Pittsburgh pneumonia agent and discuss the broader implications of dual infection regarding the epidemiology and transmission of these organisms. MATERIAL AND METHODS Isolation of Organisms. Clinical specimens were inoculated onto routine media, buffered charcoal yeast extract  and semiselective differential BCYE agar medium containing 1 yg/ml of vancomycin, 50 units/ml of polymyxin B, 0.001 percent bromthymol blue, and 0.001 percent bromcresol purple [ 111. The addition of dyes to the media allowed presumptive identi-
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SIMULTANEOUS LEGIONNAIRES’ AND PITTSBURGH PNEUMONIA-MUDER
fication of L. pneumophila and Pittsburgh pneumonia agent on the basis of colony morphology and color [ 121. Identity of isolates showing typical colony morphology, staining characteristics, and lack of growth on unsupplemented sheep blood agar was confirmed by slide agglutination  and direct fluorescent antibody testing. Direct Fluorescent Antibody Testing. Direct fluorescent antibody testing of isolates and clinical specimens was performed for L. pneumophila serogroups 1 to 6 and Pittsburgh pneumonia agent with reagents supplied by the Center for Disease Control [ 141.
Serologic Testing. Testing of patient serum samples was performed by the enzyme-linked immunosorbent assay for antibodies to L. pneumophila serogroups 1 to 6 and Pittsburgh pneumonia agent [ 151.
RESULTS Patients. Pneumonia developed in the hospital in six patients, and one became ill four days following a brief admission for an unrelated problem. In all cases, pneumonia was characterized by acute pulmonary infiltrate with cough and/or fever. Evidence of dual infection with L. pneumophila and Pittsburgh pneumonia agent was present in all patients (Table I). One patient has been described previously [ 161. All patients were hospitalized at the Pittsburgh Veterans Administration Hospital between August 1980 and February 1982. Duration of hospitalization prior to onset ranged from two to 5 1 days. Patient characteristics are summarized in Table I. Clinical Illness. The clinical presentation was not distinctive. Six patients had fever (range 101 to 104’F), and one patient had rigors at the onset of pneumonia. Four patients had cough with purulent sputum. One patient had undergone intubation for mechanical ventilatory assistance prior to the onset of pneumonia. Three patients complained of chest pain, with one patient describing the pain as pleuritic. Two patients were confused and disoriented. An additional patient was noted to be obtunded at a time when his serum sodium level was 117 meq/liter. Virus-like prodrome was observed in one patient, and diarrhea was observed in two patients. Leukocyte counts ranged from 6,200 to 43,900/mm3. Two patients had hyponatremia, with serum sodium levels of 117 to 126 meq/liter. In six patients, hepatic enzyme levels were measured during the course of the illness; modest elevations of serum transaminases (serum glutamic-pyruvic transaminase 53 to 94 mu/ml, and serum glutamic oxaloacetic transaminase 29 to 94 mu/ml) were seen in four. One patient had acute hyperbilirubinemia (serum bilirubin 15 mg/dl) that appeared shortly after the administration of rifampin and resolved following discontinuation of the drug. All patients had new pneumonic infiltrates on chest x-ray films during the course of the illness. Unilateral and bilateral infiltrates were seen; neither cavitation nor nodule formation was observed.
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Diagnosis of Legionnaires’ and Pittsburgh Pneumonia. Results of microbiologic and immunologic tests are summarized in Table I. All patients had both L. pneumophila and Pittsburgh pneumonia agent demonstrated in clinical specimens by culture, direct fluorescent antibody testing, or both (Table I). L. pneumophila was isolated by culture in four patients (lung tissue in two and respiratory secretions in two). L. pneumophila was demonstrated by direct fluorescent antibody stain in five patients (lung tissue in three, sputum in one, and both sputum and lung tissue in one). Pittsburgh pneumonia agent was cultured from the lung tissue of two patients. Pittsburgh pneumonia agent was visualized by direct fluorescent antibody stain in all seven patients (lung tissue in two, sputum in four, and both in one). Histopathologically, lung tissue showed acute bronchopneumonia in all four patients in whom autopsy was carried out. In three patients, both L. pneumophila and Pittsburgh pneumonia agent were visualized by direct fluorescent antibody stain in areas of histopathologic involvement. One patient (Patient 7) had only L. pneumophila detected in lung tissue at autopsy despite demonstration of Pittsburgh pneumonia agent in sputum. However, he had received 12 days of erythromycin therapy prior to death. Serum samples for convalescent serologic study were obtained in four patients who survived at least two weeks. Two patients showed a greater than fourfold rise in titers to both L. pneumophila and Pittsburgh pneumonia agent. One patient showed no serologic response at two weeks to either organism. One other patient showed seroconversion to L. pneumophila but not to Pittsburgh pneumonia agent at 12 weeks. Therapy and Owtcome. The diagnosis of Legionella infection was made during the hospital course in five patients. All five patients were treated with erythromycin; one patient received rifampin in addition. Despite therapy, two patients died from pneumonia; erythromycin therapy was instituted within 24 hours of the clinical onset of pneumonia in these two patients. One patient appeared to survive the pneumonia but died of other causes. In two patients, the diagnosis of Legionella infection was made from postmortem lung specimens. One patient had received cephalothin and gentamicin, and the other had received cephradine. COMMENTS The laboratory evidence demonstrating dual infection in these patients is strong. In individual patients, however, demonstration of infection by both organisms was not obtained with all three diagnostic methods (culture, direct fluorescent antibody testing, and serologic study). Two patients died without antemortem suspicion of Legionella infection, and dual infection was recognized on postmortem culture of lung tissue; thus, not all available methods were applied. In three patients,
62 M Lung cancer, cardiomy-
FlWWslXrlt Antlbsdy Culture Tsstisg Culture
Sputum Lung tissue Sputum
In two separate specimens. + Pneumonia was resolving when patient died of myocardial
61 M Metastatic prostate cancer 60 M Coronary artery disease
Steroids, cytotoxic drugs, radiation therapy Steroids
60 M Chronic alcoholism, diabetes mellitus
opathy 47 M Pulmonary sarcoidosis
58 M Carcinoma of larynx
58 M Cirrhosis
Duration of HospHaHzation pirn to Pneumonia (days)
Patients with Simultaneous Legionnaires’ Disease and Pittsburgh Pneumonia
Age/ patient Sex
2 weeks 1:1280
Acute I:80 4 weeks 1:1.280 15 weeks 1:1,280 58 weeks 0 Acute 0 3 weeks 1:160 12 weeks 1:160 Acute 0
2 weeks 1:320
Acute 1:80 4 weeks 1:320 15 weeks 1:1,280 58 weeks 1:10,240 Acute 0 3 weeks 0 12 weeks 0 Acute 1:80
Serologic Titers Pittsburgh L. pneumophila Agent
Agsst mrsct Fluorescent Antibody T=tfng
Cephalothin, gentamicin Cephradine
SIMULTANEOUS LEGIONNAIRES’ AND PITTSBURGH PNEUMONIA-MUDER
Year Reported 1976 1977
Historical Parallels between Discovery of Legionella pneumophila and Pittsburgh Pneumonia Agent Plttsburgh Pneumonia Agent
Leaionella pneimophila Undiagnosed pneumonia at the American Legion Convention  L. pneumophila isolated from lung tissue [I] L. pneumophila recognized as a cause of nosocomial pneumonia i? renal transplantation patients 
Undiagnosed nosocomial pneumonia in renal transplantation patients  Isolation of L. pneumophila from hospital 1980 Pittsburgh pneumonia agent isolated from shower  lung tissue  1982 Demonstration of widespread colonization of Isolation of Pittsburgh pneumonia agent the hospital water system by L. from hospital shower  pneumophila  Demonstration of colonization of the hospital water system by Pittsburgh Pneumonia Agent [ lo] Report of seven cases of pneumonia with simultaneous isolation of L. pneumophila and 1983 Pittsburgh pneumonia agent (present report)
neither organism was isolated by culture, although direct fluorescent antibody stain revealed the organisms. In two other patients, L. pneumophila was isolated but not Pittsburgh pneumonia agent. This may have occurred for a number of reasons. L. pneumophila is fastidious and is easily overgrow? by commensal airway flora. Pittsburgh pneumonia agent is even more fastidious in its growth habits [6, lo]. We have previously established that culture using transtracheal aspiration specimens is the most sensitive diagnostic test for Legionella infections in our institution [ 171, and none of the specimens from culture-negative patients were collected using this method. In one patient, erythromycin therapy probably contributed to our inability to isolate either organism, despite demonstration of both by direct fluorescent antibody testing of sputum and lung tissue. All isolates of L. pneumophila and Pittsburgh pneumonia agent were tested with direct fluorescent antibody conjugate against both organisms to rule out the possibility of cross-reactions, although such crossreactions among Legionella species have not been reported [ 18,191. Repeated testing in our laboratory has also failed to establish evidence of cross-reactivity. Although cross-reactivity has been reported between L. pneumophila and non-Legionella bacteria such as Bacteroides fragilis and Pseudomonas fluorescens [ 14,201, we have not observed this phenomenon in our institution in a previous study of bacterial isolates taken from sputum specimens (Vu VL, unpublished data).
serum samples were obtained in four
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patients; in two, diagnostic increases in antibody titer to both L. pneumophila and Pittsburgh pneumonia agent were observed. In one patient, convalescent serum specimens were drawn at two weeks after onset of pneumonia. No seroconversion to either species was observed, perhaps because seroconversion to L. pneumophila frequently requires four or more weeks; the rate of seroconversion at three weeks has been estimated in one study to be as low as 59 percent [ 2 11. One patient showed seroconversion to L. pneumophila but not Pittsburgh pneumonia agent; however, the sensitivity of and time required for seroconversion to Pittsburgh pneumonia agent are not precisely known. We and others have noted occasional cases in which results of direct fluorescent antibody testing or culture failed to match the species for which an antibody increase could be demonstrated. For example, Pittsburgh pneumonia agent might be isolated from a clinical specimen, yet the patient may show antibody response to L. pneumophila or to more than one Legionella species . This has raised the possibility of serologic cross-reactivity among species within the genus Legionella, since cross-reactions have been documented after experimental infection in animals [ 181. This report suggests that an alternative explanation exists, namely, the possibility of simultaneous infection by two Legionella species. During the period August 1980 through January 1982, 39 cases of nosocomial Legionnaires’ pneumonia and 26 cases of Pittsburgh pneumonia were diagnosed at the Pittsburgh Veterans Administration Hospital. Thus,
17.9 percent (seven of 39) cases of Legionnaires’ disease and 26.9 percent (seven of 26) cases of Pittsburgh pneumonia represented cases of simultaneous infection. It would appear that, at this institution, not only are L. pneumophila and Pittsburgh pneumonia agent common causes of nosocomial pneumonia, but also simultaneous infection with both organisms is not rare. Although the number of patients with dual infection is too small for meaningful comparison with those having other nosocomial pneumonias, we were nevertheless impressed by both the degree of underlying debility and the high mortality rate in the seven patients described herein. Perhaps these patients were unusually susceptible to infection by both pathogens because of the severity of underlying disease. The possibility of synergistic pathogenicity between the two species of Legionella remains to be explored. The evolution of current knowledge of L. pneumophila and Pittsburgh pneumonia agent has followed a strikingly parallel course (Table II). L. pneumophila was recovered from lung tissue after a dramatic outbreak of pneumonia for which no cause could initially be found . It was recognized as an organism that could not be placed in any existing taxonomic classification . Similarly, Pittsburgh pneumonia agent was discovered in lung tissue during the investigation of etiologically undiagnosed pneumonia [ 21 and also recognized as a previously uncharacterized species . The role of L. pneumophila as a nosocomial pathogen was appreciated after an outbreak in renal transplantation patients ; Pittsburgh pneumonia agent was also first recognized as a nosocomial pathogen in real transplant recipients [8,9]. A number of risk factors, notably im-
&Dade JE, Shepard CC, Redus MA, et al: Legionnaires’ disease: isolation of a bacterium and demonstration of its role in other respiratory diseases. N Engl J Med 1977; 297: 1197-1203. Pasculle AW, Myerowitz RL. Rinaldo CR: New bacterial agent of pneumonia isolated from renal transplant recipients. Lancet 1979; II: 58-61. Cordes LG, Wilkinson HW, Gorman GW, Fikes BJ, Fraser DW: Atypical Legionella-like organisms: fastidious water associated bacteria pathogenic for man. Lancet 1979; II: 927-930. Lewallen K, McKinney RM, Brenner DJ, et al: A newly identified bacterium phenotypicaliy resembling but genetically distinct from Legionella pneumophila: an isolate in a case of pneumonia. Ann Intern Med 1979; 91: 831-834. liebert GA, Steigerwalt AG, Brenner DJ: Legionella micdadei species nova: classification of a third species of Legionella associated with human pneumonia. Curr Microbial 1980; 3: 255-258. Pasculle AW, Feeley JC, Gibson RJ, et al: Pittsburgh pneumonia agent: direct isolation from human lung tissue. J Infect Dis 1980; 141: 727-732.
munosuppression and cigarette smoking, have been described for L. pneumophila [26,27]; the same predisposing factors have been demonstrated for Pittsburgh pneumonia agent . The isolation of L. pneumophila from hospital showers  was followed by the discovery that the organism is capable of widespread, persistent colonization of water distribution systems . Recently, Pittsburgh pneumonia agent was isolated from a hospital shower  and is now also known to be capable of widespread colonization of the water distribution system [lo]. L. pneumophila and Pittsburgh pneumonia agent show a number of microbiologic similarities. Both organisms are poorly staining gram-negative rods that fail to grow in routine culture media such as blood agar and require specialized media for isolation [ 1,6]. The organisms share flagellar antigens [ 181. Furthermore, both organisms are susceptible in vitro to erythromycin and rifampin , an unusual pattern for gram-negative organisms. Given the similarities in microbiologic characteristics, predisposing host factors, and environmental reservoirs, this report of dual infection by both organisms seems to be a fitting culmination of the evolutionary parallel of L. pneumophila and Pittsburgh pneumonia agent. There are, at least, two important clinical implications to be drawn from this report (1) Seroconversion to more than one Legionella species may be due to multiple infection as well as to serologic cross-reactivity. (2) The mode of transmission for L. pneumophila and Pittsburgh pneumonia agent remains uncertain; given these seven cases, it is likely that the mode of transmission is identical for both pathogens.
Garrity GM, Brown A. Vickers RM: Tatlockia and Fluoribacter: two new genera of organisms resembling Legionella pneumophila. Int J Syst Bact 1980; 30: 609-619. Myerowitz RL, Pasculle AW, Dowling JN, et al: Opportunistic lung infection due to “Pittsburgh pneumonia agent.” N Engl J Med 1979; 301:953-958. Rogers BH, Donowitz GR, Walker GK, Harding SA, Sande MA: Opportunistic pneumonia: a clinico-pathological study of five cases caused by an unidentified acid-fast bacterium. N Engl J Med 1979; 301: 959-961. Stout J, Yu VL, Vickers RM, Shonnard J: Potable water supply as the hospital reservoir for Pittsburgh pneumonia agent. Lancet 1982; I: 471-472. Gsrrity GM, Elder EM, Davis B, Vickers RM, Brown A: Serologic and genotypic diversity among serogroup &--reacting environmental Legionella isolates. J Clin Microbial 1982; 15: 646-653. Vickers RM, Brown A, Garrity GM: Dye-containing buffered charcoal yeast extract medium for differentiation of members of the family Legionellaceae. J Clin Microbial 1981; 13: 380-382. Wilkinson HW, Fikes BJ: Slide agglutination test for sero-
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grouping Legionella pneumophila and atypical Legionella-like organisms. J Clin Microbial 1960; 11: 99. Cherry WD, Pittman B, Harris PP, et al: Detection of Legionnaires’ disease bacteria by direct immunofluorescent staining. J Clin Microbial 1976; 6: 329-336. Farshy CE, Klein GC, Feeley JC: Detection of antibodies for the Legionnaires’ disease organism by microagglutination and micro-enzyme-linked immunosorbent assay tests. J Clin Microbial 1976; 7: 327-331. Brown A, Shonnard JW, Geyer SJ, et al: Coincident infection with Legionnaires’ disease bacterium and Pittsburgh pneumonia agent. Lancet 1961; II: 1041-1042. Zuravleff JJ, Rihs J, Shonnard J, Elder E: Sensitivity of diagnostic tests for Legionnaires’ disease: a reappraisal (abstr C39). In: Program of the 62nd Annual Meeting, American Society for Microbiology, March 7 to 12, 1962, Atlanta, Georgia. Hebert GA, Moss CW, McDougal LK. Bozeman FM, McKinney RM, Brenner DJ: The Rickettsia-like organisms TATLDCK (1943) and HEBA (1939): bacteria phenotypically similar to but genetically distinct from Legionella pneumophila and the WIGA bacterium. Ann Intern Med 1960: 92: 45-52. Cordes LG, Myerowitz RL, Pasculle AW, et.al: Legionella micdadei: (Pittsburgh pneumonia agent): direct fluorescent antibody examination of infected human lung tissue and characterization of clinical isolates. J Clin Microbial 1961; 13: 720-722. Edelstein PH, McKinney RM, Meyer RD, et al: Immunologic diagnosis of Legionnaires’ disease: cross-reactions with anaerobic and microaerophilic organisms and infections caused by them. J Infect Dis 1960; 141: 652-655. Edelstein PH, Meyer RD, Finegold SH: Laboratory diagnosis of Legionnaires’ disease. Am Rev Respir Dis 1960; 121: 317-327. Donegan EA, Deal MM, Melanephy MC, et al: Primary isoiation of a new strain of the TATLDCKlPittsburgh pneumonia agent (Legionella micdadei). West J Med 1961; 134:
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364-369. Fraser DW, Tsai TR, Drenstein W, et al: Legionnaires’ disease: description of an epidemic of pneumonia. N Engl J Med 1977; 297: 1169-l 197. Brenner DJ, Steigerwalt AG, f&Dade JE: Classification of the Legionnaires’ disease bacterium: Legionella pneumophila, genus novum, species nova, of the family Legionellaceae, familia nova. Ann Intern Med 1979; 90: 656-656. Bock BV, Edelstein PH, Snyder KM, et al: Legionnaires’ disease in renal transplant patients. Lancet 1976; I: 410413. Broome CV, Goings SAJ, Thacker SB, et al: The Vermont epidemic of Legionnaires’ disease. Ann Intern Med 1979; 90: 573-577. Haley CE, Cohen ML, Halter J, Meyer RD: Nosocomial Legionnaires’ disease: a continuing commcn-source epidemic at the Wadsworth Medical Center. Ann Intern Med 1979; 90: 563-566. Muder RR, Yu VL, Zuravleff JJ: Pneumonia due to the Pittsburgh pneumonia agent: new clinical perspective with a review of the literature. Medicine 1963; in press. Tobin J D’H, Dunnill MS, French M, et al: Legionnaires’ disease in a transplant unit: isolation of the causative agent from shower baths. Lancet 1960; II: 116-121. Stout J, Yu VL, Vickers RM, et al: Ubiquitousness of Legionalla pneumophila in the water supply of a hospital with endemic .Legionnaires’ disease. N Engl J Med 1962, 306: 466468. Brown A, Yu VL, Magnussen MH, Vickers RM, Garrity GM. Elder EM: Isolation of Pittsburgh pneumonia agent from a hospital shower. Appl Environ Microbial 1982; 43: 725726. Pasculle AW, Dowling JN, Weyant RS, et al: Susceptibility of Pittsburgh pneumonia agent (Legionella micdadei) and other newly recognized members of the genus Legionella to nineteen antimicrobial agents. Antimicrob Agents Chemother 1981; 18: 403-408.