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Background: Drug-resistant tuberculosis (TB) is associated with higher mortality rates in patients with human immunodeficiency virus (HIV). In Mexico, the number of deaths due to TB among the HIV-positive population has tripled in recent years. Methods: Ninety-three <i>Mycobacterium tuberculosis</i> strains isolated from the same number of HIV-infected patients treated in a public hospital in Mexico City were studied to determine the drug resistance to first- and second-line anti-TB drugs and to identify the mutations associated with the resistance. Results: Of the 93 patients, 82.7% were new TB cases, 86% were male, and 73% had extrapulmonary TB. Most patients (94%) with a CD4 T-lymphocyte count <350 cells/mm³ were associated with extrapulmonary TB (<i>p</i> <0.0001), whilst most patients (78%) with a CD4 T-lymphocyte count >350 cells/mm³ were associated with pulmonary TB (<i>p</i> = 0.0011). Eighty-two strains were pan-susceptible, four mono-resistant, four poly-resistant, two multidrug-resistant, and one was extensively drug-resistant. In the rifampicin-resistant strains, <i>rpoB</i> S531L was the mutation most frequently identified, whereas the <i>inhA</i> C15T and <i>katG</i> S315T1 mutations were present in isoniazid-resistant strains. The extensively drug-resistant strain also contained the mutation <i>gyrA</i> D94A. Conclusions: These data highlight the need to promptly diagnose the drug resistance of <i>M. tuberculosis</i> among all HIV-infected patients by systematically offering access to first- and second-line drug susceptibility testing and to tailor the treatment regimen based on the resistance patterns to reduce the number of deaths in HIV-infected patients.