Azlocillin

Mycobacterium avium-intracellulare complex (MAC) is a ubiquitous environmental microorganism whose pathogenicity ranges from innocuous colonization to disease, in immunocompetent as well as immunocompromised individuals. We sought to determine the clinical significance of MAC in sputum cultures of patients with pulmonary tuberculosis (TB). A retrospective analysis between January 1994 and March 1995 at Bellevue Hospital Center revealed both Mycobactenum tuberculosis and MAC in 35 patients (11% of all patients with TB). Of 27 patients reviewed, 52% were HIV-1 infected (median CD4 + 25 cells per microliter). Radiographic manifestations in patients with TB and MAC were similar to those seen in patients with TB alone. Both mycobacteria were cultured primarily from respiratory sources. M tuberculosis was usually cultured first or concurrent with MAC, and in nearly all cases, both species were recovered within 2 months of each other. Most patients improved clinically, bacteriologically, and radiographically with standard antituberculous therapy, except those with advanced AIDS, multidrug-resistant TB (MDR-TB), or disseminated MAC. We conclude that recovery of MAC in sputum is common in patients with pulmonary TB, regardless of HIV-1 infection, MDR-TB, or other clinical, bacteriologic, or radiographic attributes. MAC cultivation in most of these patients likely represents transient colonization, and in most cases is not clinically significant.

(CHEST 1997; 111:142-47)

Key words: M avium; M tuberculosis

Abbreviations: MAC=Mycobacterium avium- intracellilare complex; MDR-TB=multidrug-resistant tuberculosis; MGIT=mycobacterial growth indicator tube; OADC=oleic acid, albumin, dextrose, and catalase; TB=tuberculosis

Mycobacterium avium-intracellulare complex (MAC) is the most common systemic bacterial pathogen in AIDS. MAC incidence increases as immune function wanes, occurring in up to half of AIDS patients at autopsy.[1-3] Although MAC is detected in only 15 to 25% of AIDS patients antemortem,[1] up to 60% of patients with MAC in sputum or stool cultures may develop disseminated MAC within 1 year.[3] MAC is also an increasingly recognized pulmonary pathogen in nonimmunocompromised individuals, even without known preexisting pulmonary disease.[4-6] Concurrent with the recent tuberculosis (TB) epidemic, we have commonly observed the isolation of MAC in the sputum of patients with cultures positive for Mycobacterium tuberculosis. In contrast to M tuberculosis, in which recovery from any bodily site signifies disease, the finding of MAC in nonsterile sites usually represents colonization. The coexistence of these two mycobacteria may reflect MAC infecting damaged respiratory airways, similar to the role of bacterial species in bronchiectasis or chronic bronchitis, or it may be present only as a saprophyte. Although guidelines exist that try to distinguish MAC colonization from infection,[7,8] this distinction often is not readily apparent.

Our objectives were to determine the incidence of MAC cultivation from the sputum of patients with pulmonary TB, to evaluate whether certain patients with TB are in some way predisposed to coinfection with MAC, and to ascertain the clinical significance of MAC cultivation in patients with TB. To obtain this information, we identified patients with isolates of both M tuberculosis and MAC retrospectively and reviewed their clinical, radiographic, and microbiologic characteristics from the time of diagnosis through the completion of antituberculous therapy.

MATERIALS AND METHODS

Patient Study Group

A list of all patients with both M tuberculosis and MAC cultured from January 1, 1994 to March 31, 1995 was obtained from the microbiologic records at Bellevue Hospital Center in New York City. Patients were included in this study if they had at least one positive culture for M tuberculosis from a pulmonary source and two or more sputum cultures of MAC on at least two separate occasions. Medical records were reviewed to obtain the following information: age, gender, race, usage of alcohol, tobacco, illicit drug injection, and domiciliary status, as well as HIV-1 infection as determined by enzyme-linked immunosorbent assay and Western blot analysis, and CD4+ lymphocyte counts using flow cytometry. The clinical presentation and treatment regimens were also analyzed. Microbiologic data included ascertainment of the initial mycobacterial isolate, the sequence and timing of recovery of M tuberculosis and MAC, the culture sources, and the M tuberculosis drug-susceptibility patterns. Chest radiographs were examined to determine radiographic manifestations. Clinical outcome was determined by review of inpatient and outpatient medical records of Bellevue Hospital Center and the New York City Department of Health. Clinical improvement was defined as the resolution of respiratory and constitutional symptoms believed to be due to mycobacterial infection. Bacteriologic improvement was defined as two or more consecutive smear and culture-negative specimens on at least two separate occasions, without subsequent positive smears or cultures. Follow-up information was obtained for up to 24 months after initiation of treatment for TB.

Specimen Processing

Sputum induction was performed by having patients rinse their mouths with sterile water and then inhaling nebulized sterile 3% saline solution for 10 to 15 min or until the onset of sputum expectoration. All patient material was collected in sterile containers for processing.

All specimens were digested and decontaminated with 2% sodium hydroxide, 2.9% sodium citrate, and 5% n-acetyl cysteine for 15 min, and the reaction was terminated by the addition of an equal volume of 0.9% saline solution. The specimens were then centrifuged at 5,000 g for 15 min, decanted, and the sediment was used to inoculate media and prepare smears. Prior to October 1994, all cultures were inoculated into a system (Seph-Check AFB System; Becton-Dickinson; Cockeysville, Md), which is a sealed quadriphasic culture system. This system consists of a lower chamber filled with liquid media (Middlebrook and Cohn 7H9) supplemented with a combination of oleic acid, albumin, dextrose, and catalase (OADC) and an antibiotic supplement containing amphotericin, azlocillin, colistin, and trimethoprim, and an upper chamber with a paddle with three areas coated with agar (Middlebrook and Cohn 7H10), an agar-based variant of Lowenstein-Jensen media, and chocolate agar. The inoculum was added to the liquid media and incubated at 37[Degrees]C. The units were examined daily for growth, and negative units were inverted to seed the solid media in the upper chamber. After October 1994. the cultures were inoculated to solid media and a mycobacterial growth indicator tube (MGIT) (Becton-Dickinson). These tubes contain Middlebrook and Cohn 7H9 media supplemented with OADC and polymyxin, amphotericin B, nalidixic acid, trimethoprim, and azlocillin (PAN-TA) and have an indicator in the bottom of the tube that fluoresces as the level of oxygen in the tube decreases. These were examined daily on an ultraviolet transilluminator to detect growth. Solid media were examined weekly for 8 weeks.

Cultures showing signs of growth as turbidity in the liquid media, colonies on the solid media, or fluorescence in the MGIT tubes were verified by performing a Kinyoun acid-fast stain on the growth. Those cultures positive for mycobacterial growth were then screened for both M tuberculosis and MAC by DNA-RNA hybridization with the M tuberculosis and MAC system (AccuProbe Culture Confirmation System; Gen-Probe; San Diego, Calif). Susceptibility studies were performed using a modification of the method of proportions.[9] Isolates of M tuberculosis were tested for susceptibility to a battery of ten antimy- cobacterial agents, including isoniazid, rifampin, pyrazinamide, ethambutol, streptomycin, kanamyein, ethionamide, rifabutin, ciprofloxacin, and amikacin. MAC susceptibility was not routinely performed.

Statistics

Fisher's Exact Test was used to compare clinical findings between HIV-positive and HIV-negative patients.

RESULTS

During the study period, 295 patients had positive cultures for only M tuberculosis, 534 patients had positive cultures for only MAC, and 35 patients had positive cultures for both M tuberculosis and MAC. Medical records were available for 33 of the 35 patients with positive respiratory cultures of both M tuberculosis and MAC, but were not available for two patients who were treated at another hospital. Nineteen patients had cultures of MAC on Septi-Check and 14 patients had cultures of MAC on MGIT. Twenty-seven patients with pulmonary TB had two or more sputum cultures positive for MAC on at least two separate occasions. Six patients with TB had only one sputum culture positive for MAC and were excluded from further study.

Most of these 27 patients were young (mean age, 44 years) and were men (81%) (Table 1). HIV-1 infection was present in slightly more than half of the patients (14/27, 52%). One patient denied HIV risk factors but refused testing. Most patients were Hispanic or African-American (85%) and had a high prevalence of homelessness and drug use, including alcohol, smoking, and IV drugs. No patient reported chronic respiratory symptoms, although several patients (both with and without HIV-1 infection) had prior respiratory infections.

(*) Percentage of patients with both M tuberculosis and MAC.

Symptoms, mycobacteriology, and HIV-1 status are shown in Table 2. The median CD4+ cell count in HIV-1-infected patients was 25 cells per microliter (range, 10 to 460 cells per microliter; mean, 109 [+ or -] 153 cells per microliter). Constitutional symptoms (fever, sweats, weight loss, malaise) were present in 13 of 14 HIV-1-infected patients (93%), compared to 8 of 12 (67%) without HIV-1 infection (p = NS). Respiratory symptoms (cough, dyspnea, pleuritic pain, hemoptysis) were present in 9 of 14 HIV-infected patients (64%), compared to 11 of 12 (92%) of those without HIV-1 infection (p = NS).

Table 2--Symptoms, Mycobacteriology, and Clinical Outcome of Patients With M tuberculosis and MAC by HIV Status(*)

Both M tuberculosis and MAC were cultured primarily from respiratory sources. Interestingly, we cultured M tuberculosis from nonrespiratory sites more commonly than MAC, regardless of HIV-1 status. This may reflect the fact that M tuberculosis is a more common extrapulmonary pathogen, while MAC is commonly found in the setting of colonization, especially in the respiratory and GI tracts. Also, nonrespiratory mycobacterial cultures were not routinely performed unless patients were symptomatic; thus, these sites were likely underrepresented. Three patients in whom MAC was cultured from nonrespiratory sources (stool and blood) had AIDS and two with disseminated MAC died soon after diagnosis, despite treatment for both M tuberculosis and MAC.

Although the long-term prognosis in patients with M tuberculosis and MAC was not evaluated, nearly all patients in this study improved clinically, bacteriologically, and radiographically with standard antituberculous therapy. Only those patients with advanced AIDS, MDR-TB, or disseminated MAC had a poor outcome. Of the four known deaths in this group, all were in patients with advanced AIDS, plus disseminated TB and MAC, disseminated cytomegalovirus, neutropenia, or hepatorenal syndrome. Although four patients did not have documented clearance of MAC from sputum, each had clinical and radiographic improvement. Following standard antituberculous therapy, there were no known treatment failures, disease recurrences, or positive mycobacterial cultures for up to 24 months of follow-up.

Isolation of MAC from a normally sterile, nonpulmonary site definitively establishes infection, but the presence of MAC in sputum alone does not establish the etiology in patients with pulmonary disease.[2,7,13] In individuals with chronic lung disease (in whom MAC has been classically described), the pathogenicity of MAC may range from benign colonization to chronic, acute, or subacute infection.[8] In one retrospective review of nearly 500 patients with respiratory cultures of MAC, the authors believed that half of the isolates represented colonization on clinical grounds alone.[13] The other half were cultured from elderly white women with bronchiectasis, solitary nodules that were tuberculomas, and in less than 5% of the cases, cavitary nodular lung disease mimicking pulmonary TB.

The pathogenicity of MAC in patients with HIV-1 infection can be especially difficult to assess. In AIDS patients with disseminated MAC, chest radiographs may be normal, lung tissue damage is typically absent, and other pulmonary pathogens may coexist at autopsy.[1,14,15] Most HIV-1-infected patients with sputum cultures of MAC alone have no clinical or radiographic evidence of pulmonary disease, and that which is present is usually attributable to another etiology.[2,16,17] In a study of 32 AIDS patients with postmortem evidence of disseminated MAC, although 34% had MAC isolated from lung tissue, only 13% had microscopic evidence of pulmonary involvement, and each of these patients had coexisting pulmonary conditions sufficient to cause the observed lung abnormality.[1] Thus, MAC appears to be an uncommon cause of respiratory disease in patients with AIDS.[2,3]

In our series, most patients had signs and symptoms consistent with TB, and most patients improved with standard antituberculous treatment. This is consistent with the concept that MAC is more likely to be a saprophyte rather than a true pathogen, and supports the idea that in most patients with current or previously treated M tuberculosis, MAC is clinically insignificant.[7,8,18] HIV-1 infection in patients with M tuberculosis and MAC appeared to influence mortality in 4 patients, although this probably is a function of the HIV-1 infection rather than a consequence of true MAC disease.

The finding of MAC in the sputum of patients with M tuberculosis may be due to a combination of an increase in the prevalence of MAC[19] and a predisposition of patients with TB to become colonized with MAC. Although respiratory tract cultures of MAC do not often precede MAC dissemination in AIDS patients, the isolation of MAC in respiratory secretions has been shown to have a high positive predictive value for progression to dissemination.[3,17,20] Hence, it is possible that a number of HIV-1-infected patients in our series may progress to disseminated MAC infection. Conversely, MAC cultivation in these patients may merely represent transient colonization in relation to pulmonary TB, as we believe occurs in those without HIV-1 infection.

Our study is limited by several factors. Because this was a retrospective analysis, follow-up cultures were obtained at the discretion of the treating physicians. Although we limited our inclusion of patients to those with a minimum number of culture samples both before and after treatment, four patients did not have documented bacteriologic clearance, despite clinical and radiographic resolution. Also, any diagnostic testing or treatment (for TB) that patients received outside of New York City was not known to us. Finally, although MAC has been a subject of growing interest in AIDS patients and nonimmunocompromised individuals, the incidence of MAC in the sputum of healthy, asymptomatic individuals is not known.

In a patient whose clinical and radiographic findings are compatible with TB, but with sputum cultures positive only for MAC, we recommend an aggressive approach to rule out TB. In patients with sputum cultures negative for TB, we routinely pursue bronchoscopy with transbronchial biopsy, transbronchial lymph node aspiration, and analysis of samples by polymerase chain reaction. If all cultures remain negative for M tuberculosis, a trial of empiric therapy for TB may still be given, and if there is a response, a full course of empiric therapy should be administered. If there is no improvement with antituberculous therapy, and MAC is consistently cultivated, empiric treatment for MAC may then be given.

We conclude that the coexistence of M tuberculosis and MAC is common, regardless of HIV-1 infection or other demonstrated characteristics. MAC cultivation from the sputum of most patients with pulmonary TB likely represents colonization, and in most cases it is transient and not clinically significant. There are rare circumstances in which M tuberculosis and MAC may simultaneously cause disease, however, and specific treatment for both mycobacteria is then required.[7] In all instances, the pathogenicity of MAC is best determined by a combination of clinical, radiographic, and pathologic criteria.[7,13,21]

ACKNOWLEDGMENT: The authors thank Natalie Little for expert editorial assistance and H. William Harris, MD, for critical review of the manuscript.

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(*) From the Departments of Medicine (Drs. Epstein, Aranda, and Rom) and Pathology (Mr. Bonk and Dr. Hanna), NYU Medical Center, and the Division of Pulmonary and Critical Care Medicine, Bellevue Chest Service, New York. Supported by GCRC grant RR00096, NIH AI35233, HL51494, and CDC CCU210075.

Manuscript received March 28, 1996; revision accepted June 6. Reprint requests: Dr. Rom, NYU Medical Center, Bellevue Chest Service, 550 First Avenue, New York, NY 10016

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