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Rapid and accurate laboratory detection and identification of Mycobacterium tuberculosis, particularly multidrug-resistant strains, is critical to both public health control measures and patient management. The authors surveyed microbiology laboratories to evaluate whether their methods met national guidelines. As needed, laboratories received individualized recommendations for improvement. The laboratories were resurveyed a year later to assess changes in methods. Current guidelines recommend fluorochrome acid-fast smears, broth cultures, identification by nucleic acid probe or BACTEC-NAP, and BACTEC primary susceptibility panels, which should include pyrazinamide. Of 27 laboratories performing acid-fast smears, 15 used fluorochrome methods. Six of 16 laboratories performing mycobacterial cultures used broth media. Of six laboratories performing species identification, five used nucleic acid probes or BACTEC-NAP. Of five laboratories evaluating drug sensitivity, two used BACTEC and two included pyrazinamide in their protocols. Overall, 24 (89%) laboratories needed improvements; a year later, 16 (67%) of those had altered their methods or made definite plans to do so. Survey results suggest that health departments can facilitate improvements in laboratory testing for pathogens of public health importance.