Corynebacterium equi Infection Complicating Neoplastic Disease RICHARD BERG, M.D., HERMAN CHMEL, M.D., JOAN MAYO, B.A., AND DONALD ARMSTRONG, M.D.

THE ISOLATION of a Corynebacterium, commonly reported as a "diphtheroid," is usually regarded as normal flora or a "contaminant" with no clinical significance. This may be true; however, occasionally these organisms can cause serious and even fatal infections. We recently observed two patients, one with reticulum cell sarcoma and the other with Hodgkin's disease, both of whom had pulmonary abscesses and septicemia with Corynebacterium equi. These are the fourth and fifth instances of an invasive human infection with this organism, and as in the previous three cases, the hosts were immunosuppressed. The clinical settings and characteristics of the organism are reported. Received March 25, 1976; received revised manuscript July 16, 1976; accepted for publication July 16, 1976. Supported by the American Cancer Society Clinical Investigation Grant #26. Dr. Chmel was supported by a training program for Cancer Teaching and Research, National Cancer Institute No. CA 05110. Address reprint requests to Dr. Armstrong; Memorial Hospital, 1275 York Avenue, New York, New York 10021.

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From the Infectious Disease Service, the Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York, and the Cornell University Medical College, New York, New York

Report of Two Cases Case I. A 52-year-old woman was admitted to another hospital in September 1973 for evaluation of lymphadenopathy, which revealed reticulum cell sarcoma. Past medical history was significant for two episodes of hemolytic anemia, initially treated with splenectomy in 1949 and with corticosteroids in 1961. In October 1973, the patient was treated with Cytoxan, vincristine, and prednisone, which treatment was continued for a total of eight cycles until May 1974. On May 28, a lymphangiogram revealed persistent lymph nodal enlargement, and a chest x-ray demonstrated a right hilar mass and pathologic rib fractures on the left. The patient then received radiation therapy to the abdomen and left lateral chest wall, which she tolerated well, until July 8, 1974. when fever, nausea, vomiting, weakness, and pancytopenia developed. By July 15, the patient had daily fevers, the temperatures as high as 39 C. with a cough productive of brownish sputum, which on culture revealed normal mouth flora. On July 21, treatment with prednisone, 60 mg per day, was begun. On August 9, the patient was admitted to Memorial Sloan-Kettering Cancer Center for evaluation of fever. There was no history of travel outside metropolitan New York City. The patient had no direct contact with animals. Positivefindingson physical examination were intermittent rales and decreased breath sounds in the left lower lung field. Examination of the heart revealed a regular tachycardia without a murmur. Chest x-ray showed a left pleural reaction that had been present on June 10. Results of laboratory studies were: leukocyte count, 4,900/mm3 with 90% polymorphonuclear leukocytes; hemoglobin 10.9 g/dl; platelet count 52,000/mm3; blood urea nitrogen 11.7 mg/dl; total protein 5.8 g/dl; albumin 2.1 g/dl; total bilirubin 1.5 mg/dl; alkaline phosphatase 308 international units; serum glutamic oxaloacetic acid transaminase 160 Karmen units; positive direct and indirect Coomb's tests; quantitative immunoglobulin IgG 2,100 mg/dl, IgM 160 mg/dl, IgA 215 mg/dl. The patient's hospital course was characterized by daily fevers with temperatures as high as 40 C, accompanied by chills. A

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Berg, Richard, Chmel, Herman, Mayo, Joan, and Armstrong, Donald: Corynebacterium equi infection complicating neoplastic disease. Am J Clin Pathol 68: 73-77. 1977. Corynebacterium equi, a soil-residing diphtheroid pathogenic in horses, swine and cows, caused pulmonary infection with bacteremia in two patients with lymphomas. Both patients were being treated with immunosuppressive therapy, as were the patients in three previously reported human cases. Unless certain characteristics of these organisms are recognized, they may be regarded as normal flora or contaminating diphtheroids. They could also be mistaken for other grampositive rods, such as Bacillus species, Listeria monocytogenes, or Erysipelothrix insidiosa. C. equi isolates have usually been sensitive to erythromycin, tetracycline, gentamicin, and carbenicillin. One of the patients reported here was successfully treated with erythromycin, tetracycline, and surgery. (Key words: Corynebacterium equi; Lymphoma; Leukemia; Immunosuppressed patient; Opportunistic infection.)

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FIG. 2. Roentgenogram of the chest, Case 2, before operation showing infiltrates throughout the right lung, but primarily in the right upper lobe.

blood culture drawn on August 10 yielded an aerobic grampositive rod, which was later identified as Corynebacterium equi. The organism was sensitive to erythromycin, tetracycline, gentamicin and kanamycin, and resistant to penicillin, ampicillin, oxacillin, cephalothin and lincomycin, using the Kirby-Bauer technic.1 The results of tube-dilution tests done subsequently are recorded in Table 2. Corynebacterium equi was cultured aerobically from six blood cultures obtained from August 10 through August 29. On August 16, treatment with cephalothin, 2 g every 6 hours (130 mg/kg/day) and gentamicin, 60 mg every 6 hours (4 mg/kg/day), intravenously, was begun. A chest film revealed a fluid level in the left lower lung field, and a peripheral wedge-like infiltrate in the left lower lung field (Fig. 1). On August 20, gentamicin was increased to 5 mg/kg/day and cephalothin to 12 g per day (200 mg/kg/day), intravenously. On August 19 and 22, thoracenteses were performed, revealing bloody pleural fluid that contained C. equi. On August 22, a chest tube was placed in the left thoracic space. On August 26, the patient's serum was shown to be bactericidal for her organism at a dilution of 1:8, while she was receiving gentamicin and cephalothin. A repeat serum immunoglobulin determination August 30 revealed IgG 470 mg/dl, IgM 37 mg/dl, IgA 80 mg/dl. The remainder of the patient's hospital course was characterized by deterioration of pulmonary status, cardiac arrhythmias, hypotension, azotemia, and gastrointestinal bleeding. The patient died September 1. Permission for autopsy was refused. Case 2. A 47-year-old white woman who had had Hodgkin's disease since 1970 was admitted to Memorial Sloan-Kettering Cancer Center for evaluation of fever and dry cough. From 1970 through 1973, the patient had been treated with many chemotherapeutic agents and radiation therapy, and she was symptomfree by the end of 1973. In June 1974, the Hodgkin's disease recurred, manifested by fever, pelvic mass and lymph nodal enlargement, and the patient was treated with Cytoxan, vincristine, procarbazine, prednisone, and levamisole, with partial regression. In October 1974, treatment with adriamycin, vinblastine,

and bleomycin was started. In November the bleomycin was discontinued and prednisone started. During the same month, the patient complained of cough and dyspnea on exertion. These symptoms slowly progressed, and the patient was admitted to another hospital on January 16, 1975. Significant history revealed that she lived in a rural area next to a field where horses had grazed. In addition, the patient was fond of outdoor gardening and used dried cow's manure as a fertilizer in her garden. On admission, she complained of fever and cough. A chest film revealed an infiltrate in the right upper lobe (Fig. 2). The temperature was 39 C. The patient had a pustular lesion on her forehead and another at the angle of her mouth on the right side of her face. The initial leukocyte count was 14,900/mm3, with 95% polymorphonuclear leukocytes. A percutaneous-transthoracic lung biopsy yielded a pure culture of Corynebacterium equi. The same organism was isolated from blood cultures and scraping of the cutaneous lesions. The patient was treated with erythromycin, and a right upper lobectomy was performed. Pathologic study of the submitted lung tissue revealed acute necrotizing pneumonia with abscess formation on a background of chronic inflammatory changes withfibrosis.Special stains showed a homogeneous collection of gram-positive coccobacillary organisms. Cultures yielded C. equi in pure culture. The organism was sensitive to erythromycin, tetracycline, gentamicin and kanamycin, and resistant to penicillin, ampicillin, oxacillin, cephalothin and lincomycin by the disk method, results of tube dilutions done subsequently are recorded in Table 2. Postoperatively, the patient's temperature returned to normal, and she was discharged February 22, receiving erythromycin, 2 g per day, orally. Two weeks after discharge, the patient stopped taking erythromycin, with the subsequent development of fevers, with temperatures as high as 38.5 C, and dry cough. At the end of February, a draining right thigh sinus developed; the facial lesion also began to drain. On March 25, a sore throat developed and the patient was admitted to Memorial Sloan-Kettering Cancer Center on March 27. On physical examination, she was afebrile; examination of the throat showed diffuse maculopapular lesions,

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FIG. 1. Roentgenogram of the chest, Case 1, revealing a wedgeshaped infiltrate and fluid in the left lower lung field. There are also peripheral infiltrates in the right lower lung field.

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Table I. Biochemical Reactions of Cultures of Corynebacterium equi Test

Result

Catalase Oxidase Kligler iron agar Hydrogen sulfide (lead acetate paper) Christensen's urea Indole Ornithine decarboxylase Lysine decarboxylase Arginine dihydrolase Nitrate reduction Simmons' citrate Phenylalanine deaminase Gelatin hydrolysis Deoxyribonuclease Bile esculin Growth in 6.5% NaCI ONPG* Dextrose Maltose Lactose Mannitol Arabinose Sucrose Adonitol Inositol Rhamnose Raffinose Salicin Sorbitol Xylose Trehalose

Positive Negative Alkaline/alkaline Positive Positive Negative Negative Negative Negative Positive Negative Negative Negative Positive Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative Negative

* ONPG — 0-Nilrophenyl-beta-D-g;ilacto-pyranoside.

Table 2. Concentrations of Antibiotics Inhibiting Growth of Corynebacterium equi (MIC) MIC

Penicillin, units/ml Ampicillin, /xg/ml Tetracycline, /xg/ml Chloramphenicol, |itg/ml Erythromycin, /xg/ml Kanamycin, /xg/ml Gentamicin. /ng/ml Cephalothin, /ug/ml Carbenicillin. ^.g/ml Colistin, /ig/ml

Isolate A*

Isolate Bt

Result

64 16 1.0 2.0 0.12 16.0 1.0 64 16 128

32 16 2.0 4.0 0.12 16.0 0.5 32 16 128

Resistant Resistant Sensitive Sensitive Sensitive Sensitive Sensitive Resistant Sensitive Resistant

* Isolate of Corynebacterium equi from Patient 1. t Isolate of Coiynebacleritim equi from Patient 2. gentamicin was stopped and tetracycline re-started at 1 g per day, orally. The patient continued to feel improved subjectively and was discharged on May 7, receiving erythromycin, 2 g per day. and tetracycline, 1 g per day, orally. Erythromycin was continued, and the patient was doing well when last seen in August 1975. Between April 30 and July 9, 1975, on the presumption that this might aid in the treatment of her infection or Hodgkin's disease, her attending physician gave the patient 5 units of irradiated lymphocytes, donated by her two sons. Bacteriologic Results Corynebacterium equi is an aerobic gram-positive coccobacillus that is pleomorphic when grown in liquid medium. On solid medium (brain-heart infusion) the organisms were coccoid one hour, bacilloid three hours, and coccoid 24 hours after inoculation. They did not grow in thioglycollate broth or on MacConkey's agar. The organisms grew well at 37 C, were nonhemolytic, produced a pink pigment, were mucoid, and when inoculated at the top of a brainheart infusion slant, the growth ran down the slant. The biochemical reactions of both organisms are summarized in Table 1. They were catalase-positive, oxidase-negative, and gave no reaction with the sugars tested. They were nonmotile and did not form spores, which helped to differentiate them from a Bacillus species. Table 2 summarizes the minimal inhibitory concentrations (MIC), which were determined by the serial tube-dilution method. 3 Each isolate of C. equi from each patient was sensitive to erythromycin, tetracycline, gentamicin, and kanamycin, and resistant to penicillin and ampicillin at blood levels achieved by the usual doses. Discussion Corynebacterium equi is found in soil. In horses, swine and cattle, it may cause suppuration of the lungs,

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and indirect laryngoscopy revealed a white membrane. Examination of the skin revealed pustules on the forehead and the right angle of the mouth, subcutaneous masses over the back and both lower extremities, and a draining sinus from the right thigh. Laboratory findings included: leukocyte count 6,100/mm3, with 66% polymorphonuclear leukocytes and 34% lymphocytes: elevated alkaline phosphatase, 135 units; immunoelectrophoresis normal, with quantitative immunoglobulin IgG 1,350 mg/dl, IgA 84 mg/dl, and IgM 115 mg/dl. There was no reaction to dinitro chlorobenzene (DNCB), mumps, PPD and streptokinase/streptodornase (SK/SD) skin test reagents. Cultures of material from all skin lesions, the throat, and the subcutaneous masses were performed: the latter proved to be abscesses. The cultures all yielded Corynebacterium equi, sensitive to erythromycin, tetracycline, gentamicin and kanamycin; resistant to cephalothin, ampicillin and penicillin. The chest roentgenogram on admission revealed nodular densities in the right upper lung below apical pleural thickening (no change from the roentgenogram obtained in February 1975). Treatment with erythromycin, 2 g per day, was begun on March 28. Serum bactericidal levels determined after the patient had received erythromycin for two days revealed levels of 1:16 5 minutes before the next dose, and 1:32 30 minutes after the last dose of erythromycin. On March 30, tetracycline, 1 g per day, orally, was added, because one isolate of C. equi from a cutaneous lesion was more sensitive to tetracycline than to erythromycin. On April 7, gentamicin, 4 mg/kg/day, was started intravenously, in four equally divided doses, and the tetracycline discontinued. The patient appeared to be doing well clinically, with alleviation of her cutaneous lesions and pulmonary symptoms, and on April 21,

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Corynebacterium parvum has been used as an immunopotentiating vaccine in the treatment of neoplastic disease. We did not notice any effect of C. equi on the progress of the neoplasms in the patients reported here. Three previous reports of human infection with this organism have been published. The first case involved a 29-year-old man with plasma cell hepatitis receiving treatment with 60 mg prednisone per day and 6-mercaptopurine. A pulmonary abscess and an infected lymph node due to C. equi developed; both resolved in response to therapy with erythromycin. The patient had a history of cleaning pens in stockyards that housed cattle, sheep, and swine. 4 The second case was that of a 39-year-old man who had

lymphosarcoma, treated with prednisone, cytotoxic chemotherapy, splenectomy and radiation. The patient was seen because of pulmonary infiltrates. Bronchial washings and blood cultures yielded C. equi. He was treated with ampicillin and his condition improved, but he had a relapse two months later, with blood cultures positive for C. equi. Autopsy disclosed numerous pulmonary abscesses, which on culture yielded Enterococcus and Aspergillus fumigatus. This patient had spent several days working at a country fair and owned four horses. 12 The third case 15 was that of a 35-year-old woman who had had a renal transplant for progressive uremia due to proliferative glomerulonephritis. Four years after transplantation, the patient had a left-upper-lobe pneumonia which, over a two-month period, cavitated, necessitating a thoracotomy that resulted in a left upper lobectomy. Culture of the lung tissue yielded C. equi. The authors stated that she subsequently remained well. Patient 1 presented in this paper was an immunosuppressed host with reticulum cell sarcoma and no contact with a known reservoir of C. equi; Patient 2 was also an immunosuppressed host, with Hodgkin's disease and a history of exposure to horses and cow manure. Pulmonary abscesses developed in all five patients with C. equi infections while they were receiving cytotoxic drugs and prednisone. Three patients were bacteremic. Two patients had had splenectomies, and a third patient had an enlarged spleen with a background of cirrhosis. Penicillin, cephalothin, and erythromycin are the agents of choice for the treatment of most diphtheroid infections, 8 which have included life-threatening illnesses such as endocarditis, meningitis, and osteomyelitis. 7 ' 8 The antibiotic sensitivities of Corynebacterium equi reported in the literature have been variable; the organisms reported here were sensitive to erythromycin, tetracycline, kanamycin, and gentamicin, and resistant to penicillin and ampicillin. With the latter two antibiotics, however, inhibitory or microbiocidal levels should be achievable by administering higher doses than usual, and monitoring blood levels. Of the five patients reviewed in this paper, one was cured with erythromycin, two patients died although their organisms were sensitive to the antibiotics used, one patient was treated by surgical excision, and one patient, having completed a sixmonth course of treatment with erythromycin in August 1975, is doing well. The role of the lymphocyte transfusion in this patient's clinical response is uncertain. Of the three patients infected with C. equi reported in the literature, one had bacteremia recorded. The two patients reported here had bacteremia.

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lymph nodes and endometrium. 2 , 9 1 0 1 4 1 6 The portal of entry of infection is felt to be the respiratory tract. 10 The organism has been isolated in 34 cultures from human specimens as identified and recorded by the Center for Disease Control through 1970.5 Corynebacterium equi belongs to the diphtheroids, which are commonly present on the skin and in the pharynx, urethra and vagina. Isolation of diphtheroids from clinical specimens usually reflects normal flora or contamination, and therefore has no clinical significance, although they can cause fatal infections. 7,8 Corynebacterium equi could be confused with diphtheroids and considered normal flora or a contaminant. Corynebacterium equi has been reported to be acidfast,13 (our isolates were), and there is some question whether the organism is more closely related to, and therefore should be classified with, the mycobacteria. One of our isolates grew on LowensteinJensen medium, and due to its acid-fast reaction, could have at first been confused with a mycobacterium. C. equi may be confused with certain strains of Streptococci, which become rod-shaped under adverse conditions 11 ; Listeria monocytogenes, a motile, catalase-positive, gram-positive rod causing betahemolysis on sheep blood agar; or Erysipelothrix insidiosa, a nonmotile, catalase-negative, gram-positive rod that is nonhemolytic. Our first patient was initially thought to be infected with a Bacillus species because of the staining characteristics of the organism, clinical presentation, and rapid onset of pleural effusion with a wedge-shaped infiltrate.6 However, unlike Bacillus species, C. equi does not form spores and is nonmotile. C. equi is, however, similar to Bacillus species in that it is sensitive to chloramphenicol, tetracycline, and gentamicin. Final identification of the organism Corynebacterium equi was made by the Center for Disease Control, Atlanta, Georgia.

A.J.C.F. • July 1977

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cal Microbiology. American Society for Microbiology, Bethesda, Maryland, 1970, pp 93 6. Ihde DC, Armstrong D: Clinical spectrum of infection due to Bacillus species. Am J Med 55:839-845, 1973 7. Johnson WD, Kaye D: Serious infections caused by diphtheroids. Ann NY Acad Sci 174:568-576, 1970 8. Kaplan K, Weinstein L: Diphtheroid infections of man. Ann Intern Med 70:919-929, 1969 9. Karlson AG, Moses HE, Feldman WH: Corynebacterium equi in the submaxillary lymph nodes of swine. J Infect Dis Acknowledgments. Dr. Monroe Dowling of Memorial Sloan67:243-251, 1970 Kettering Cancer Center offered the authors the opportunity to 10. Knight HD: Corynebacterium infections in the horse: Problems see and study the first patient; Doctors Harry Linhardt, FishkiU of prevention. J Am Vet Med Assoc 155:446-452, 1969 Medical Group, FishkiU, New York, and Charles W. Young, 11. Lamanna C: The non-life cycle explanation of the diphtheroid Memorial Sloan-Kettering Cancer Center, referred the second paStreptococcus from endocarditis. J Bacterid 47:327-334, tient to us. 1944 12. March JC, von Graevenitz A: Recurrent Corynebacterium References equi infection in lymphoma. Cancer 32:147-149, 1973 1. Bauer AW, Kirby WMM, Sherris JC, et al: Antibiotic suscepti13. Rogosa M, Cummins RA, Leiliott RA, et al: Coryneform bility testing by a standard single disc method. Am J Clin group of bacteria, Bergey's Manual of Determinative BacPathol 45:493-496, 1966 teriology. Eighth edition. Baltimore, Williams and Wilkins, 2. Dimock WW, Edwards PR: Corynebacterium equi pneumopia 1974, pp 606-607 14. Smith JE: Corynebacterium species as animal pathogens. in foals. J Amer Vet Med Assoc 79:809, 1931 J Appl Bacteriol 29:119-130, 1966 3. Gavan TL: Broth dilution methods, Antimicrobial Susceptibility 15. Williams GD, Glanigan WJ, Campbell GS: Surgical manageTesting. American Society of Clinical Pathologists, Chicago, ment of localized thoracic infections in immunosuppressed Illinois, 1971, pp 105-126 patients. Ann Thorac Surg 12:471-480, 1971 4. Golub B, Falk G, Spink WW: Lung abscess due to Corynebacterium equi: Report of first human infection. Ann Intern 16. Woodroofe GM: Studies on strains of Corynebacterium equi isolated from pigs. Aust J Exp Biol Med Sci 28:399-409, Med 66:1174-1177, 1967 1950 5. Hermann GJ, Weaver RE: Corynebacterium. Manual for Clini-

The first patient had a sustained bacteremia suggestive of endocarditis, but died without an autopsy. The other patient had no evidence of endocarditis. Limited experience with this organism in human infections reveals a primarily pulmonary, acute-tosubacute illness, which may respond to appropriate therapy, even in the immunosuppressed host.

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Corynebacterium equi infection complicating neoplastic disease.

Corynebacterium equi Infection Complicating Neoplastic Disease RICHARD BERG, M.D., HERMAN CHMEL, M.D., JOAN MAYO, B.A., AND DONALD ARMSTRONG, M.D. TH...
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