Ultrastructure and genetic evidence of “Candidatus Rickettsia andeanae” within the Gulf Coast tick

Gulf Coast ticks, Amblyomma maculatum, occur in most southeastern United States, and are the vector of Rickettsia parkeri, a mild human rickettsiosis similar to Rocky Mountain spotted fever. A novel spotted fever group rickettsia (SFGR), and potential endosymbiont, ″Candidatus Rickettsia andeanae″ was detected in A. maculatum and is still poorly understood. We report 15%(105/698) of A. maculatum infected singly with R. parkeri, 1.4%(10/698) with ″Ca. R. andeanae″ only, and additionally 1.9%(13/698) ticks co-infected with both species, the first evidence of co-infection in A. maculatum. Identification of ″Ca. R. andeanae″ in singly and co-infected A. maculatum led us to investigate the biology of this novel SFGR. To our knowledge, no morphological study of ″Ca. R. andeanae″ has been performed, nor studies to ascertain which tick tissues are infected. Transmission electron microscopy (TEM) offers the ability to closely evaluate internal structure of the tick and associated rickettsiae, including tissue tropism of the rickettsiae. We found evidence of rickettsiae in adult A. maculatum that were positive for the ″Ca. R. andeanae″ by PCR. Infected ticks contained rod-like bacteria of small size (approximately 0.3x0.9um), often with a halo zone (electron-lucent layer around the organism), and a trilaminar cell wall, all consistent with SFGR morphology. Female ticks contained organisms suggestive of ″Ca. R. andeanae″ in granular salivary glands, foregut, Malpighian tubules, nerve and reproductive tissue. Interestingly, bacteria were less common in male ticks, but present in agranular salivary glands. These findings contribute to the current understanding of this recently identified rickettsia in A. maculatum.