Background: Pertussis continues to be an important health issue in Vietnam despite infant vaccination programs. In Vietnam, the incidence rates of pertussis per 100,000 population rose from 0.09 in 2014 to 0.33 in 2015 and to 0.58 in 2017. Macrolides, especially erythromycin, are the treatment of choice. However, erythromycin-resistant cases, caused by transition at A2047G position in 23S rRNA, have been reported in the region. Few data are available on antimicrobial resistance in Bordetella pertussis to guide treatment in Vietnam. We report antimicrobial susceptibility of the circulating strains in southern Vietnam during 2015–2017. Methods: Tracheal aspirates from 263 suspected pertussis cases were subject to multiplex real-time PCR to identify B. pertussis and Bordetella spp. Samples were cultured on Regan Lowe agar with 10% sheep blood containing cephalexin (40 µg/mL) and incubated at 37°C for 10 days. The antimicrobial susceptibilities to erythromycin, azithromycin, clarithromycin, and trimethoprim/sulfamethoxazole were determined using the disc diffusion method (CLSI-2017) on Regan Lowe and Mueller Hinton agar. Erythromycin minimum inhibitory concentrations (MICs) were determined using an E-test. The results were recorded after days 3 and 7 of incubation. Sequencing of the 23S rRNA gene was performed to detect mutations conferring macrolide resistance. Results: Of 263 cases, 119 were positive for B. pertussis (45.2%) by real-time PCR, and 15 of 263 strains (5.7%) were successfully cultured. All 15 isolates were susceptible to macrolides and no heterogeneous phenotype was recorded after 7 days; erythromycin MICs were ≤0.094 µg/mL (Fig. 1). We observed no difference in results generated on Regan Lowe and Mueller Hinton media. However, for testing trimethoprim/sulfamethoxazole, results on were superior, as those on Regan Lowe media were unclear. Sequencing of 23S rRNA identified no mutations known to confer macrolide resistance. Conclusions: None of 15 B. pertussis isolates tested were nonsusceptible to erythromycin and macrolides. Similarly, no mutation at the erythromycin-binding site in the 23S rRNA gene was identified. The low isolation rate of B. pertussis by culture means that few positive specimens were tested for antimicrobial susceptibility. To overcome this limitation, detection of resistance directly from clinical specimens needs to be investigated. Ongoing screening for B. pertussis and antimicrobial susceptibility is recommended to support efforts to control the spread of this respiratory tract infection agent.