SETTING: All non-private hospitals in Malawi that registered TB cases in 2001, during which there was a bus service for transporting sputum specimens to the Central Reference Laboratory (CRL) for mycobacterial culture and drug sensitivity testing (CDST). OBJECTIVES: To determine the performance of the system of collecting and processing sputum specimens from patients with recurrent smear-positive pulmonary TB through to CDST. DESIGN: Structured interviews with TB Officers, and retrospective data collection using TB and laboratory registers. RESULTS: There were 964 patients with recurrent smear-positive PTB. TB Officers took responsibility for collecting and transporting sputum to the CRL, and 73% reported using the bus service. Sputum specimens from 384 (40%) patients arrived at the CRL. Of these, 40% were found to have negative concentrated smears at the CRL, and 36% of specimen sets arriving at CRL were successfully cultured for DST. Most specimens had been collected after the start of anti-tuberculosis treatment. Although delays in collection adversely affected culture, only 43% of specimen sets collected on or before the first day of treatment yielded Mycobacterium tuberculosis. CONCLUSION: Problems were identified at all stages of the system and strategies to remedy these are being put in place.