title: High prevalence of spotted fever group rickettsiae in Amblyomma variegatum from Uganda and their identification using sizes of intergenic spacers.
abstract: The spotted fever group (SFG) rickettsiae are obligate intracellular bacteria transmitted by ticks that cause several tick-borne rickettsioses in humans worldwide. This study was intended to determine the prevalence of SFG rickettsiae in Amblyomma variegatum from 7 districts across Uganda. In addition to sequencing of gltA and ompA genes, identification of Rickettsia species based on the sizes of highly variable intergenic spacers, namely, dksA-xerC, mppA-purC, and rpmE-tRNA(fMet) was carried out. Application of multiplex PCR for simultaneous amplification of 3 spacers combined with capillary electrophoresis separation allowed simple, accurate, and high-throughput fragment sizing with considerable time and cost savings. Rickettsia genus-specific real-time PCR detected 136 positives out of 140 samples, giving an overall prevalence of 97.1%. Most samples (n=113) had a size combination of 225, 195, and 341 bp for dksA-xerC, mppA-purC, and rpmE-tRNA(fMet), respectively, which was identical to that of R. africae, a causative agent of African tick bite fever. In addition, several samples had size variants in either dksA-xerC or rpmE-tRNA(fMet). Nonetheless, the partial sequences of gltA and ompA genes of samples of all size combinations showed the greatest similarity to R. africae (99.3-100% for gltA and 98.1-100% for ompA). Given these results, it is highly possible that the tested ticks were infected with R. africae or closely related species. This is a first report on molecular genetic detection of R. africae and its high endemicity in Uganda. Clinicians in this country should be aware of this pathogen as a cause of non-malarial febrile illness. This study provided a starting point for the development of Rickettsia species identification based on the sizes of intergenic spacers. The procedure is simple, rapid, and cost-effective to perform; hence it might be particularly well suited for preliminary species identification in epidemiological investigations. The results may be more detailed and reliable when simultaneous sequencing analysis is performed.
authors: Division of Collaboration and Education, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan; Division of Bioinformatics, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan.
authors: Ryo
authors: R
authors: Nakao
authors: Division of Collaboration and Education, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan.
authors: Yongjin
authors: Y
authors: Qiu
authors: Division of Bioinformatics, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan.
authors: Manabu
authors: M
authors: Igarashi
authors: Division of Collaboration and Education, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan; National Livestock Resources Research Institute (NaLIRRI), Tororo, Uganda.
authors: Joseph W
authors: JW
authors: Magona
authors: Division of Collaboration and Education, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan.
authors: Lijia
authors: L
authors: Zhou
authors: Division of Bioinformatics, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan.
authors: Kimihito
authors: K
authors: Ito
authors: Division of Collaboration and Education, Hokkaido University Research Center for Zoonosis Control, Kita 20, Nishi 10, Kita-ku, Sapporo, Hokkaido 001-0020, Japan. Electronic address: sugimoto@czc.hokudai.ac.jp.
authors: Chihiro
authors: C
authors: Sugimoto
mesh_terms: D000818:Animals; D001096:Arachnid Vectors / Q000382:microbiology*; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002417:Cattle; D002418:Cattle Diseases / Q000382:microbiology / Q000469:parasitology / Q000635:transmission*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D021901:DNA, Intergenic / Q000737:chemistry / Q000235:genetics; D005260:Female; D026863:Ixodidae / Q000382:microbiology*; D008297:Male; D008969:Molecular Sequence Data; D060885:Multiplex Polymerase Chain Reaction / Q000662:veterinary; D015995:Prevalence; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary; D012281:Rickettsia / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D012282:Rickettsia Infections / Q000453:epidemiology / Q000635:transmission / Q000662:veterinary; D017422:Sequence Analysis, DNA / Q000662:veterinary; D013984:Tick Infestations / Q000382:microbiology / Q000469:parasitology / Q000662:veterinary*; D017282:Tick-Borne Diseases / Q000453:epidemiology / Q000382:microbiology / Q000662:veterinary*; D013997:Time Factors; D014454:Uganda / Q000453:epidemiology; D015047:Zoonoses
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D021901:DNA, Intergenic
keywords: African tick bite fever; Amblyomma variegatum; Intergenic spacer; Rickettsia africae; Uganda; Zoonosis