VECTOR-BORNE AND ZOONOTIC DISEASES Volume 8, Number 1, 2008 © Mary Ann Liebert, Inc. DOI: 10.1089/vbz.2007.0138 Rickettsia amblyommii Infecting Amblyomma americanum Larvae ELLEN Y. STROMDAHL, 1 MARY A. VINCE, 1 PEGGY M. BILLINGSLEY, 2 NICOLE A. DOBBS, 3 and PHILLIP C. WILLIAMSON 2 ABSTRACT Polymerase chain reaction analysis of Amblyomma americanum adults, nymphs, and larvae from Aberdeen Prov- ing Ground, MD (APG), revealed a very high prevalence of a spotted fever group (SFG) rickettsia. Restriction frag- ment length polymorphism (RFLP) and sequence analysis identified “Rickettsia amblyommii.” This organism is not yet described or well studied, and its pathogenicity is unknown; however, investigations of the organism are warranted because of its high prevalence in A. americanum. This tick is extremely abundant at military training facilities in the south, central, and Mid-Atlantic United States, and many soldiers experience multiple concurrent tick bites. Bites by R. amblyommii-infected A. americanum may account for rates of SFG rickettsia seropositivity that are higher than reported rates of Rocky Mountain spotted fever (RMSF) cases from the same location. Sero- conversion to SFG rickettsia following bites of A. americanum may suggest that R. amblyommii is infectious in humans. Subclinical infection in the numerous A. americanum tick bite victims could contaminate donated blood and compromise immunodeficient recipients. Detection of R. amblyommii in questing A. americanum larvae sug- gests transovarial transmission. The absence of R. rickettsii, the agent of RMSF, in A. americanum may be due to transovarial interference by R. amblyommii. The likelihood of pathogen transmission by larvae is magnified by their habit of mass attack. The very small size of the larvae is also a risk factor for pathogen transmission. High R. amblyommii prevalence in populations of A. americanum presage co-infection with other A. americanum-borne pathogens. A. americanum nymphs and adults from APG were found to be co-infected with R. amblyommii and Borrelia lonestari, Ehrlichia chaffeensis and Ehrlichia ewingii, respectively, and larval pools were infected with both R. amblyommii and B. lonestari. Co-infections can compound effects and complicate diagnosis of tick-borne disease. Key words: Rickettsia amblyommii—Amblyomma americanum—Prevalence—Pathogenicity—Co-infec- tion. 15 INTRODUCTION T HE TICK-BORNE DISEASE LABORATORY of the Entomological Sciences Program of the U.S. Army Center for Health Promotion and Preventive Medicine (USACHPPM) adminis- ters the Department of Defense (DOD) Human Tick Test Kit Program by providing expedient analyses of ticks removed from military per- sonnel. Large numbers of these ticks are Am- blyomma americanum, including larvae. This tick is extremely abundant at military training fa- cilities in the south, central and mid-Atlantic United States, and many soldiers experience multiple concurrent tick bites (Stromdahl et al. 2001, 2003). Polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis (RFLP) of nucleic acid preparations 1 Entomological Sciences Program, U.S. Army Center for Health Promotion and Preventive Medicine, Aberdeen Proving Ground, Maryland. 2 Department of Pathology and Human Identification, University of North Texas Health Science Center, Ft. Worth, Texas. 3 Department of Molecular Biology & Immunology, University of North Texas Health Science Center, Ft. Worth, Texas.