Survival of mycobacteria on N95 personal respirators.

OBJECTIVES: The overall aim of this study was to investigate the survival and possible growth of Mycobacterium tuberculosis simulant bacteria on respirator filters. METHODS: Mycobacterium smegmatis was used as a biochemical simulant for M tuberculosis. Bacterial survival was tested on National Institute for Occupational Safety and Health-certified N95 respirators from three manufacturers. The first experiments simulated one-time respirator use and subsequent storage for 1 to 9 days under ideal conditions for the growth of mycobacteria: 37 degrees C and 85% relative humidity. The bacteria were loaded on the respirator filters under three different nutritional conditions: in the absence of nutrients; in the presence of human saliva (simulating conditions when the respirator is worn); and in the presence of nutrient broth (for ideal growth potential). The subsequent experiments simulated respirator wear for 2 hours under medium work-load conditions at a breathing rate of 56 L/min. RESULTS: It was found that M smegmatis did not grow on the tested respirators, even when the respirators were stored at temperature, humidity, and nutrition conditions most favorable for microbial growth. However, these bacteria could survive on respirators for up to 3 days during storage. The culturability of M smegmatis was not affected by airflow that simulated the breathing rate associated with medium work-load conditions for 2 hours. CONCLUSIONS: This study shows that M tuberculosis surrogate bacteria collected on a respirator are not able to grow and are able to survive only in ideal (ie, not clinically relevant) conditions. Based on these experiments, we conclude that M tuberculosis is unlikely ever to become an infectious problem in the air again, once it is removed by a respirator.

Analysis of airborne actinomycete spores with fluorogenic substrates.

The reactions between seven fluorogenic substrates and different groups of enzymes, esterases, lipases, phosphatases, and dehydrogenases, were studied in a search for a new method for the detection of actinomycete spores. Fluorescence measurement was chosen as a fast and sensitive method for microbial analysis. The focus of the research was on the spores of important air contaminants: Streptomyces albus and Thermoactinomyces vulgaris. For the measurement of the enzymatic activity, the chosen fluorogenic substrate was added to a mixture of spores and nutrient media, and the resulting fluorescence was measured with a spectrofluorometer. Fluorogenic substrates were found to show enzymatic activities even for dormant spores. Comparison of the enzymatic activities of dormant spores with those of vegetative cells showed similarity of the enzymatic profiles but higher activity for vegetative cells. The increase of enzymatic activity from dormant spores to vegetative cells was not linear but fluctuating. The largest fluctuations were found after 4 to 5 h of incubation. The enzymatic activities of S. albus were 10 to 50 times lower than those of T. vulgaris, except for the dehydrogenase activity, which was seven times higher. These results indicate that analysis with fluorogenic substrates has the potential for becoming a fast and sensitive method for the enumeration and identification of airborne actinomycete spores.

Characteristics of airborne actinomycete spores.

Airborne actinomycete spores, important contaminants in occupational and residential environments, were studied with respect to their (i) release into the air, (ii) aerodynamic and physical size while airborne, and (iii) survival after collection onto agar with an impactor. Three actinomycete species were selected for the tests to exemplify the three main spore types: Streptomyces albus for arthrospores, Micromonospora halophytica for aleuriospores, and Thermoactinomyces vulgaris for endospores. The results show that the incubation conditions (temperature, time, and nutrients) needed for the development of spores for their release into air are different from the conditions that are needed for colony growth only. Additional drying of M. halophytica and T. vulgaris cultures was needed before spores could be released from the culture. The aerodynamic sizes of the spores, measured with an aerodynamic particle sizer, ranged from 0.57 (T. vulgaris) to 1.28 micron (M. halophytica). The physical sizes of the spores, when measured with a microscope and an image analysis system, were found to be smaller than previously reported in the literature. The relative recovery of the spores on agar media ranged from 0.5 (T. vulgaris) to 35% (S. albus). The results indicate that the culturability of the collected airborne actinomycete spores varies widely and is affected by several variables, such as the species and the sampling flow rate. Therefore, alternatives to commonly used cultivation methods need to be developed for the enumeration of actinomycete spores.