| Summary: | <i>Rickettsia conorii</i> is the causative agent of Mediterranean spotted fever (MSF). Misdiagnosis of MSF may occur with febrile syndromes associated with rash and thrombocytopenia, such as Crimean-Congo hemorrhagic fever (CCHF). This study aimed to determine the prevalence of <i>R. conorii</i> among serum samples obtained from 260 suspected CCHF patients with features of MSF in Iran (2018–2020). The quantitative polymerase chain reaction (qPCR) method detected three (1.15%) positive 16S rDNA <i>Rickettsia</i> spp. samples that were classified as <i>R. conorii</i> subsp. <i>conorii</i>, <i>R. conorii</i> subsp. <i>Israelensis,</i> and <i>R. helvetica</i> using the sequencing of <i>gltA</i>, <i>ompA</i>, and <i>17kDa</i> genes. Furthermore, <i>R. conorii</i> IgM antibodies presented in 38 (14.62%) patients by the enzyme-linked immunosorbent assay (ELISA) method. Out of 97 MSF patients with available paired serum samples, IgM seroconversion and a four-fold increase were observed in 14 (14.43%) and 12 (12.37%) patients, respectively. We concluded that rickettsial agents are present in Iran and may be misdiagnosed with other febrile syndromes.
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