Nosocomial Mycobacterium fortuitum colonization from a contaminated ice machine.

Between October 15 and November 18, 1985, 5 patients on a medical ward of the Albany VA Medical Center (Ward 8A) became colonized with Mycobacterium fortuitum. Because other patients in Ward 8A were at risk of developing disease with M. fortuitum, microbiologic surveillance to identify colonization in sputum was begun. By February 15, 1986, 30 colonized patients had been identified in this ward but none in another ward with a comparable patient population, which suggests a source unique to Ward 8A. Because water has been recognized as a source of opportunistic mycobacterial pathogens, we conducted a retrospective case-control study using a telephone survey questionnaire to examine a number of water exposures in 10 patients and 20 control subjects. Exposure to ice from the Ward 8A ice machine, but not to potable water, was associated with colonization with M. fortuitum. Large-volume water samples from a variety of sources were cultured for acid-fast bacilli. M. fortuitum was isolated only from the ice machine in Ward 8A. The ice machine was disconnected, and no additional patients became colonized. Although ice machines are infrequently implicated in nosocomial outbreaks, they represent a potential source for pathogens that survive or replicate in water.

Epidemiology of infection by nontuberculous mycobacteria. V. Numbers in eastern United States soils and correlation with soil characteristics.

Organisms of the Mycobacterium avium, M. intracellulare, and M. scrofulaceum (MAIS) group were recovered in significant numbers from 63 soil samples collected from the floodplains of 4 major rivers in the eastern United States: Hudson (NY), James (VA), Savannah (GA, SC), and Tombigbee (AL, MS). The mean numbers of M. avium-intracellulare, M. scrofulaceum, and MAIS-intermediate biovars recovered increased significantly from the Hudson to the James to the Tombigbee and Savannah soil samples, agreeing reasonably well with previous findings on MAIS distribution in eastern U.S. waters and with the geographic distribution of naval recruits reacting to PPD-B and PPD-G, which is higher in the southeastern United States. From the Savannah and Tombigbee rivers, soils were collected throughout their lengths. The number of total MAIS and biovars recovered from the different soil sites varied widely and showed no obvious trends. However, statistical analysis revealed a highly significant correlation between high numbers of MAIS recovered and high acidity of the soils. Other soil characteristics, such as high total organic matter, high conductivity, and reduced moisture, were only weakly, if at all, correlated with high MAIS numbers. Whereas the general distribution of MAIS in these soils of the Northeast (NY), mid-Atlantic (VA), and Southeast (GA, SC, AL, and MS) might explain the geographic distribution of the naval recruits reacting to PPD-B and PPD-G, the distribution of MAIS in soils along the stretches of the 2 southeastern rivers was not correlated with the county-by-county frequency of PPD-B reactive naval recruits in Georgia, South Carolina, Alabama, and Mississippi.

Phage-typing and drug-resistance patterns as tools in mycobacterial epidemiology.

Phage-typing by itself was not sufficient to delineate the boundaries of a mini-epidemic of tuberculosis in upstate New York. Drug-resistance patterns were needed as well. In a small upstate community, 79% of 14 isolates tested were resistant to one or more of the antituberculosis drugs. Of 15 isolates with phage types determined, 47% were type 1(13), 27% were type 7(7, 13), and 27% were type 1(7, 12, 13). By combining phage-typing and sensitivity testing, we were able to demonstrate that 4 or possibly 5 of the 7 phage-type 1(13) strains are epidemiologically related.

In vitro susceptibility of human and environmental isolates of Mycobacterium avium, M. intracellulare, and M. scrofulaceum to heavy-metal salts and oxyanions.

Because of the widespread distribution of Mycobacterium intracellulare and M. scrofulaceum in southeastern U.S. waters, the susceptibility of members of these species to heavy-metal salts and oxyanions was investigated. Isolates with abnormally high tolerance to mercuric chloride or cadmium chloride were identified.

Epidemiology of infection by nontuberculous mycobacteria. IV. Preferential aerosolization of Mycobacterium intracellulare from natural waters.

We report here the first laboratory studies simulating the conditions for natural aerosolization of Mycobacterium intracellulare and M. scrofulaceum and estimate the yields for this pathway of transfer of pathogenic mycobacteria from water to air; M. intracellulare and M. scrofulaceum were both concentrated in droplets ejected from cell suspensions of densities comparable to those found in natural freshwaters (100 to 2,000 colony-forming units per ml). The enrichment factor (defined as the concentration of cells per droplet volume divided by the concentration of cells in the bulk suspension per equivalent volume) for M. intracellulare isolates ranged from 68 to 15,000, with an average of 2,922; for M. scrofulaceum it ranged from 35 to 550, with an average of 177. One factor responsible for the greater aerosolization of M. intracellulare was their aggregation. However, after vortexing, M. intracellulare were still aerosolized more (enrichment factor, 325) than M. scrofulaceum. Increasing salt concentrations enriched the aerosolization of both species, but the number of organisms transferred from water to air did not increase proportionately because the salt decreased the droplet volume. Other waterborne pathogens such as Legionella pneumophila were also enriched and transferred from water to air, indicating that this pathway for possible infection of humans may also be significant for other respiratory diseases.

Differences in biological activity among batches of lyophilized tice Bacillus Calmette-Guérin and their association with clinical course in Stage I lung cancer.

Various measures of biological activity were studied in batches of lyophilized Tice Bacillus Calmette-Guérin (BCG) that have been used intrapleurally as adjuvant therapy in surgically resected Stage I lung cancer patients by our own group, the North American Lung Cancer Study Group, and the Ludwig Lung Cancer Study Group. The biological activities of BCG that were studied were: (a) inhibition of solid tumor growth in mice following s.c. inoculation of BCG admixed with methylcholanthrene-induced fibrosarcoma cells (MC 43); (b) protection against tumor colonization of lungs and prolongation of survival in mice pretreated intrapleurally with BCG and later inoculated i.v. with a lethal dose of MC 43 tumor cells; and (c) growth properties in vitro, including a determination of the relative amounts of growing and nongrowing material and the sensitivity of growing material to inhibition by isoniazid. Significant differences in biological activity among batches of BCG were found. Some batches of BCG protected mice against circulating tumor cells, while others did not. Batches of BCG which protected mice against circulating tumor cells were found to have greater sensitivity to isoniazid and higher viability than batches which did not elicit this antitumor activity. There were also trends of some batches of BCG outperforming others in the clinic. Cumulative disease-free interval of patients was longer with batches which protected mice against circulating tumor cells than with batches which did not protect mice against circulating tumor cells. The results of this study suggest that preclinical testing of BCG for antitumor activity may improve the efficacy of this agent in future clinical trials.

Uric acid utilization by Mycobacterium intracellulare and Mycobacterium scrofulaceum isolates.

Forty-nine human and environmental isolates of Mycobacterium intracellulare and Mycobacterium scrofulaceum were tested for their ability to grow on uric acid and a number of its degradation products. Nearly all (88 to 90%) strains used uric acid or allantoin as a sole nitrogen source; fewer (47 to 69%) used allantoate, urea, or possibly ureidoglycollate. Enzymatic activities of one representative isolate demonstrated the existence of a uric acid degradation pathway resembling that in other aerobic microorganisms.

Recent experience in the epidemiology of disease caused by atypical mycobacteria.

To obtain information on the epidemiology of mycobacteriosis, water and air samples collected along the East Coast of the United States were examined for mycobacteria. Mycobacterium avium-intracellulare-scrofulaceum (MAIS) were isolated from 25% of the water, samples, mostly those from South Carolina, Georgia, and the Gulf states. Mycobacterium kansasii and Mycobacterium marinum were not found, probably because of the detrimental effects of the NaOH used to decontaminate the samples. MAIS strains were found more often in estuaries than in fresh or ocean waters. The frequency of Mycobacterium intracellulare was relatively uniform along the entire coast, while Mycobacterium scrofulaceum predominated in the South. Only M. intracellulare was found in aerosol specimens, although both M. intracellulare and M. scrofulaceum were found in waters collected at the same sites.

Epidemiology of infection by nontuberculous Mycobacteria. III. Isolation of potentially pathogenic mycobacteria from aerosols.

Nontuberculous mycobacteria (21 isolates), biochemically similar to those that are recovered from humans, were recovered from rainwater and from natural river waters and their aerosols in the area of Richmond, Virginia. Field experiments have confirmed the existence of a natural mechanism for the transfer of significant numbers of mycobacteria from water to air. These findings support the hypothesis that aerosolization of potentially pathogenic mycobacteria from waters of the southeastern United States may be a major pathway for human infection.

Epidemiology of infection by nontuberculous mycobacteria. I. Geographic distribution in the eastern United States.

The nontuberculous mycobacterial group Mycobacterium avium-M. intracellular-M. scrofulaceum (MAIS) was isolated from 33% of the water samples collected from various aquatic environments in the southeastern United States. By contrast, only 20% of the water samples collected in the northeastern United States (New Jersey northward) yielded MAIS organisms. The most frequent recovery of MAIS organisms (37%) was from water samples with salinities from 0.1 to 1.9 g% (grams of NaCl/100 ml of sample). Other saprophytic slow- and rapid-growing nontuberculous mycobacteria were also isolated. The fewer MAIS organisms recovered from marine waters (20%) relative to those from freshwaters (37%) suggested that ocean water may not be a primary origin of these pathogens, although it still may be a source of infection. Our data implied a positive correlation between the frequency of persons reacting to MAIS antigens and the presence of these potential pathogens in the coastal region of the eastern United States.

Postulated sources of Mycobacterium intracellulare and Mycobacterium scrofulaceum infection: isolation of mycobacteria from estuaries and ocean waters.

Mycobacteria biochemically and serologically similar to those isolated from humans have been found in estuaries and ocean waters in the south-eastern United States. This is consistent with the hypothesis that aerosols from these waters are one potential source of infection with these mycobacteria.

Evaluation of mycobacteriology laboratories: the acid-fast smear.

In a proficiency testing program, slides bearing acid-fast bacilli were sent to laboratories in New York State. The most reliable results were obtained in laboratories using the Ziehl-Neelsen method. Cold staining procedures were less reliable. The auramine-rhodamine fluorescent stain, which produced many false positives, should be confirmed by the Ziehl-Neelsen procedure. The testing program described can be used to determine whether laboratories perform the acid-fast stain effectively.

Properties of a unique catalase isolated from Aspergillus niger.

The properties of an unusual catalase isolated from Aspergillus niger are reported. The enzyme, which has a molecular weight of 323,000, consists of four subunits and is highly resistant to sodium azide.

The tween opacity test as an aid in classification of mycobacteria.

The Tween opacity test can be used to differentiate (1) Mycobacterium flavescens from Mycobacterium gordonae and Mycobacterium szulgai, and (2) nonphotochromogenic strains of Mycobacterium kansaii from strains of the Mycobacterium terrae complex.

Three simple tests as an adjunct to the niacin test for the small mycobacteriology laboratory.

With the change in the management of tuberculosis, many bacteriology laboratories should be prepared to examine sputum for the presence of acid-fast bacilli. The niacin test is the most reliable test that can be performed in any mycobacteriology laboratory to differentiate Mycobacterium tuberculosis from the other acid-fast bacilli, but it is not perfect. Other tests can be used to supplement it are cord formation, growth at 24 C, and the catalase test at 68 C. Incompletely identified strains should be submitted to a reference laboratory.

A mycobacteriology proficiency testing program using simulated sputum specimens.

The New York State Department of Health proficiency testing program in mycobacteriology determines the ability of a laboratory to isolate the acid-fast bacillus present in a simulated specimen and to identify the strain. Until 1964 the specimens were autoclaved normal sputa seeded with mycobacteria. When mandatory testing was introduced by law in 1964, more specimens were needed. These have been prepared in simulated sputum bases, first skimmed milk, then granulated hog gastric mucin. The bases are seeded with Mycobacterium tuberculosis and other mycobacteria; Pseudomonas aeruginosa is added to simulate the contamination in clinical specimens. Laboratory performance in general has improved as a result of the proficiency testing and the concomitant educational program, but laboratories processing more than 200 specimens a year continue to perform best.