The genera Anaplasma, Ehrlichia, Cowdria, Neorickettsia and Wolbachia encompass a group of obligate intracellular bacteria that reside in vacuoles of eukaryotic cells and were previously placed in taxa based upon morphological, ecological, epidemiological and clinical characteristics. Recent genetic analyses of 16S rRNA genes, groESL and surface protein genes have indicated that the existing taxa designations are flawed. All 16S rRNA gene and groESL sequences deposited in GenBank prior to 2000 and selected sequences deposited thereafter were aligned and phylogenetic trees and bootstrap values were calculated using the neighbour-joining method and compared with trees generated with maximum-probability, maximum-likelihood, majority-rule consensus and parsimony methods. Supported by bootstrap probabilities of at least 54 %, 16S rRNA gene comparisons consistently clustered to yield four distinct clades characterized roughly as Anaplasma (including the Ehrlichia phagocytophila group, Ehrlichia platys and Ehrlichia bovis) with a minimum of 96 1 % similarity, Ehrlichia (including Cowdria ruminantium) with a minimum of 97 7 % similarity, Wolbachia with a minimum of 95 6 % similarity and Neorickettsia (including Ehrlichia sennetsu and Ehrlichia risticii ) with a minimum of 94 9 % similarity. Maximum similarity between clades ranged from 87 1 to 94 9 %. Insufficient differences existed among E. phagocytophila, Ehrlichia equi and the human granulocytic ehrlichiosis (HGE) agent to support separate species designations, and this group was at least 98 2 % similar to any Anaplasma species. These 16S rRNA gene analyses are strongly supported by similar groESL clades, as well as biological and antigenic characteristics. It is proposed that all members of the tribes Ehrlichieae and Wolbachieae be transferred to the family Anaplasmataceae and that the tribe structure of the family Rickettsiaceae be eliminated. The genus Anaplasma should be emended to include Anaplasma (Ehrlichia) phagocytophila comb. nov. (which also encompasses the former E. equi and the HGE agent), Anaplasma (Ehrlichia) bovis comb. nov. and Anaplasma (Ehrlichia) platys comb. nov., the genus Ehrlichia should be emended to include Ehrlichia (Cowdria) ruminantium comb. nov. and the genus Neorickettsia should be emended to include Neorickettsia (Ehrlichia) risticii comb. nov. and Neorickettsia (Ehrlichia) sennetsu comb. nov. Keywords : J. S. Dumler and others INTRODUCTIONRecent improvements in molecular technologies have significantly advanced our abilities to conduct genetic analyses and, for the first time, clearly indicated the proper phylogenetic positions of most of the fastidious bacterial species in the families Rickettsiaceae, Bartonellaceae and Anaplasmataceae in the order Rickettsiales (Woese et al., 1990 ;Weisburg et al., 1989 ; Brenner et al., 1993 ; Birtles et al., 1995). By 16S rRNA sequencing, Weisburg et al. (1989) demonstrated that Coxiella burnetii and Wolbachia persica belonged to the γ-Proteobacteria, while the remaining members of the order Ri...

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This investigation documents the presence of Rocky Mountain spotted fever in eastern Arizona, with common brown dog ticks (R. sanguineus) implicated as a vector of R. rickettsii. The broad distribution of this common tick raises concern about its potential to transmit R. rickettsii in other settings.

Small subunit rRNA sequences have been determined for representative strains of six species of the family Rickettsiaceae: Rickettsia rickettsii, Rickettsia prowazekii, Rickettsia typhi, Coxiella burnetii, Ehrlichia risticii, and Wolbachia persica. The relationships among these sequences and those of other eubacteria show that all members of the family Rickettsiaceae belong to the so-called purple bacterial phylum. The three representatives of the genus Rickettsia form a tight monophyletic cluster within the a subdivision of the purple bacteria. E. risticii also belongs to the a subdivision and shows a distant yet specific relationship to the genus Rickettsia. However, the family as a whole is not monophyletic, in that C. burnetii and W. persica are members of the -y subdivision. The former appears to show a specific, but rather distant, relationship to the genus Legionella.The order Rickettsiales (32) comprises a collection of procaryotes that share the property of intimate association with eucaryotic cells. The relationship in most cases is obligate intracellular parasitism, although a few of these bacteria have been grown in complex host-cell-free culture media. Although this order contains notorious pathogens of humans and animals, some of these bacteria cause no obvious damage to their host, and the relationship can be regarded as commensal. The Rickettsiales are clearly separate from the Chlamydiales (17), a narrowly defined group of energy-parasitizing, obligately intracellular bacteria. Certain fastidious parasites of plant vascular tissues and arthropods, sometimes referred to as rickettsialike (5), are not included in either category. Also, the highly heterogeneous group of bacteria that have established an endosymbiotic relationship with their hosts and for the most part have not been cultivated, including the hydrothermal-vent-associated symbionts (3,4,21), are viewed as distinct from the Rickettsiales (32) in the latest edition of Bergey's Manlual of Systematic Bacteriology.The order Rickettsiales contains three families: Rickettsiaceae, Bartonellaceae, and Anaplasmataceae. Our present study is confined to the family Rickettsiaceae, which is divided into three tribes: Rickettsieae, Ehrlichieae, and Wolbachieae (32). In the family Rickettsiaceae, rigorous criteria of classical taxonomy have been applied to the definition of most species and, in some but not in all cases, the genera. For example, good evidence was obtained by phenotypic analysis, DNA base ratio determinations, and DNA-DNA hybridization studies that in the genus Rickettsia, members of the typhus and spotted fever groups are related (32). However, the degree of relatedness of these microorganisms to the scrub typhus rickettsia, Rickettsia tsutsugamushi, remains unknown. Similarly, phenotypic analysis links the monocytic erhlichiae (Ehrlichia canis, Ehrlichia sennetsu, and Ehrlichia risticii) to each other (10,22,23), but their relationships to the granulocytic ehrlicheae * Corresponding author. t Present address: Gene-Trak Systems,

Morbidity and mortality caused by rickettsioses have had a major influence on military activities and public health for >2000 years. The threat posed by the rickettsioses is reviewed, focusing on the impact and epidemiology of those that have adversely influenced wartime operations and the current challenges posed by these diseases. With their uneven worldwide distribution, the discovery of drug-refractory strains of Orientia tsutsugamushi, the increased threat of their use in acts of bioterrorism, frequent deployment of troops to regions of endemicity, and exposures due to increased humanitarian missions, these diseases continue to be a threat to military personnel in the field. Effective strategies to reduce the impact of these diseases include development of effective vaccines, enhanced surveillance, and development of new safe, effective, and odorless repellants. The continuation of a proven, highly productive military infectious disease research program is essential for providing solutions to these daunting tasks.

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Two specific and sensitive polymerase chain reaction (PCR) assays were developed to detect and quantitate Orientia tsutsugamushi, the agent of scrub typhus, using a portion of the 47-kD outer membrane protein antigen/ high temperature requirement A gene as the target. A selected 47-kD protein gene primer pair amplified a 118-basepair fragment from all 26 strains of O. tsutsugamushi evaluated, but it did not produce amplicons when 17 Rickettsia and 18 less-related bacterial nucleic acid extracts were tested. Similar agent specificity for the real-time PCR assay, which used the same primers and a 31-basepair fluorescent probe, was demonstrated. This sensitive and quantitative assay determination of the content of O. tsutsugamushi nucleic acid used a plasmid containing the entire 47-kD gene from the Kato strain as a standard. Enumeration of the copies of O. tsutsugamushi DNA extracted from infected tissues from mice and monkeys following experimental infection with Orientia showed 27-5552 copies/microL of mouse blood, 14448-86012 copies/microL of mouse liver/spleen homogenate, and 3-21 copies/microL of monkey blood.

Bartonella species cause serious human infections globally, including bacillary angiomatosis, Oroya fever, trench fever, and endocarditis. We describe a patient who had fever and splenomegaly after traveling to Peru and also had bacteremia from an organism that resembled Bartonella bacilliformis, the causative agent of Oroya fever, which is endemic to Peru. However, genetic analyses revealed that this fastidious bacterium represented a previously uncultured and unnamed bartonella species, closely related to B. clarridgeiae and more distantly related to B. bacilliformis. We characterized this isolate, including its ability to cause fever and sustained bacteremia in a rhesus macaque. The route of infection and burden of human disease associated with this newly described pathogen are currently unknown.