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Species-specific differentiation of Bordetella pathogens in nasopharyngeal swabs of patients with suspected whooping cough was carried out using developed by us real-time PCR: duplex TaqMan PCR aimed for detecting B. pertussis and B. parapertussis, with primers targeting IS481, IS1001, BP0026 and GAPDH as the human internal control and two singleplex TaqMan PCR aimed for detecting B. holmesii and B. bronchiseptica with primers targeting hIS1001 and flaA specific genes. Among the 3130 throat swab analyzed, B. pertussis, B. parapertussis and B. holmesii were detected in 29.5 %, 0.9 % and 0.4 % of samples respectively. The developed algorithm proved to be a useful molecular-genetic diagnostic tool for whooping cough, established cocirculation of different species of Bordetella genus and demonstrated the predominance of B. pertussis, low rate of B. parapertussis and B. holmesii and the absence of B. bronchiseptica in the circulation in the Republic of Belarus.