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Objectives This study examined the phenomenon of heteroresistance in Mycobacterium tuberculosis clinical isolates obtained from retreated patients in Beijing, China between 2006 and 2011. Methods The iPLEX Gold assay platform was used to determine the prevalence of heteroresistance to injectable second-line drugs (amikacin, kanamycin and capreomycin) in resistant isolates. Results Heteroresistance was identified in 10.9% of 220 phenotypic amikacin-resistant isolates. Conclusions Heteroresistance was related mainly to the short duration and repeated use of amikacin and capreomycin during retreatment. These findings further our understanding of the evolution of resistance to injectable drugs used for tuberculosis treatment and help guide the rational use of injectable drugs during therapy. [ABSTRACT FROM PUBLISHER]