Diagnosis and Management of Tickborne Rickettsial Diseases: Rocky Mountain Spotted Fever and Other Spotted Fever Group Rickettsioses, Ehrlichioses, and Anaplasmosis - United States.

Tickborne rickettsial diseases continue to cause severe illness and death in otherwise healthy adults and children, despite the availability of low-cost, effective antibacterial therapy. Recognition early in the clinical course is critical because this is the period when antibacterial therapy is most effective. Early signs and symptoms of these illnesses are nonspecific or mimic other illnesses, which can make diagnosis challenging. Previously undescribed tickborne rickettsial diseases continue to be recognized, and since 2004, three additional agents have been described as causes of human disease in the United States: Rickettsia parkeri, Ehrlichia muris-like agent, and Rickettsia species 364D. This report updates the 2006 CDC recommendations on the diagnosis and management of tickborne rickettsial diseases in the United States and includes information on the practical aspects of epidemiology, clinical assessment, treatment, laboratory diagnosis, and prevention of tickborne rickettsial diseases. The CDC Rickettsial Zoonoses Branch, in consultation with external clinical and academic specialists and public health professionals, developed this report to assist health care providers and public health professionals to 1) recognize key epidemiologic features and clinical manifestations of tickborne rickettsial diseases, 2) recognize that doxycycline is the treatment of choice for suspected tickborne rickettsial diseases in adults and children, 3) understand that early empiric antibacterial therapy can prevent severe disease and death, 4) request the appropriate confirmatory diagnostic tests and understand their usefulness and limitations, and 5) report probable and confirmed cases of tickborne rickettsial diseases to public health authorities.

Prevalence and clinical presentation of Rickettsia, Coxiella, Leptospira, Bartonella and chikungunya virus infections among hospital-based febrile patients from December 2008 to November 2009 in Bangladesh.

BACKGROUND: We conducted a study to identify Rickettsia, Coxiella, Leptospira, Bartonella, and Chikungunya virus infections among febrile patients presenting at hospitals in Bangladesh. METHODS: We collected blood samples from patients at six tertiary hospitals from December 2008 to November 2009 and performed laboratory tests at the United States Centers for Disease Control and Prevention (CDC). RESULTS: Out of 720 enrolled patients, 263 (37%) were infected with Rickettsia; 132 patients had immunofluorescence antibody titer >64 against spotted fever, 63 patients against scrub typhus fever and 10 patients against typhus fever. Ten patients were identified with Coxiella. We isolated Leptospira from two patients and Bartonella from one patient. Ten patients had antibodies against Chikungunya virus. The proportion of patients who died was higher with rickettsial fever (5%) compared to those without a diagnosis of rickettsial infection (2%). None of the patients were initially diagnosed with rickettsial fever. CONCLUSIONS: Rickettsial infections are frequent yet under-recognized cause of febrile illness in Bangladesh. Clinical guidelines should be revised so that local clinicians can diagnose rickettsial infections and provide appropriate drug treatment.

High prevalence and two dominant host-specific genotypes of Coxiella burnetii in U.S. milk.

BACKGROUND: Coxiella burnetii causes Q fever in humans and Coxiellosis in animals; symptoms range from general malaise to fever, pneumonia, endocarditis and death. Livestock are a significant source of human infection as they shed C. burnetii cells in birth tissues, milk, urine and feces. Although prevalence of C. burnetii is high, few Q fever cases are reported in the U.S. and we have a limited understanding of their connectedness due to difficulties in genotyping. Here, we develop canonical SNP genotyping assays to evaluate spatial and temporal relationships among C. burnetii environmental samples and compare them across studies. Given the genotypic diversity of historical collections, we hypothesized that the current enzootic of Coxiellosis is caused by multiple circulating genotypes. We collected A) 23 milk samples from a single bovine herd, B) 134 commercial bovine and caprine milk samples from across the U.S., and C) 400 bovine and caprine samples from six milk processing plants over three years. RESULTS: We detected C. burnetii DNA in 96% of samples with no variance over time. We genotyped 88.5% of positive samples; bovine milk contained only a single genotype (ST20) and caprine milk was dominated by a second type (mostly ST8). CONCLUSIONS: The high prevalence and lack of genotypic diversity is consistent with a model of rapid spread and persistence. The segregation of genotypes between host species is indicative of species-specific adaptations or dissemination barriers and may offer insights into the relative lack of human cases and characterizing genotypes.

Diagnosis and management of Q fever--United States, 2013: recommendations from CDC and the Q Fever Working Group.

Q fever, a zoonotic disease caused by the bacterium Coxiella burnetii, can cause acute or chronic illness in humans. Transmission occurs primarily through inhalation of aerosols from contaminated soil or animal waste. No licensed vaccine is available in the United States. Because many human infections result in nonspecific or benign constitutional symptoms, establishing a diagnosis of Q fever often is challenging for clinicians. This report provides the first national recommendations issued by CDC for Q fever recognition, clinical and laboratory diagnosis, treatment, management, and reporting for health-care personnel and public health professionals. The guidelines address treatment of acute and chronic phases of Q fever illness in children, adults, and pregnant women, as well as management of occupational exposures. These recommendations will be reviewed approximately every 5 years and updated to include new published evidence.

When outgroups fail; phylogenomics of rooting the emerging pathogen, Coxiella burnetii.

Rooting phylogenies is critical for understanding evolution, yet the importance, intricacies and difficulties of rooting are often overlooked. For rooting, polymorphic characters among the group of interest (ingroup) must be compared to those of a relative (outgroup) that diverged before the last common ancestor (LCA) of the ingroup. Problems arise if an outgroup does not exist, is unknown, or is so distant that few characters are shared, in which case duplicated genes originating before the LCA can be used as proxy outgroups to root diverse phylogenies. Here, we describe a genome-wide expansion of this technique that can be used to solve problems at the other end of the evolutionary scale: where ingroup individuals are all very closely related to each other, but the next closest relative is very distant. We used shared orthologous single nucleotide polymorphisms (SNPs) from 10 whole genome sequences of Coxiella burnetii, the causative agent of Q fever in humans, to create a robust, but unrooted phylogeny. To maximize the number of characters informative about the rooting, we searched entire genomes for polymorphic duplicated regions where orthologs of each paralog could be identified so that the paralogs could be used to root the tree. Recent radiations, such as those of emerging pathogens, often pose rooting challenges due to a lack of ingroup variation and large genomic differences with known outgroups. Using a phylogenomic approach, we created a robust, rooted phylogeny for C. burnetii. [Coxiella burnetii; paralog SNPs; pathogen evolution; phylogeny; recent radiation; root; rooting using duplicated genes.].

Assessment of real-time PCR assay for detection of Rickettsia spp. and Rickettsia rickettsii in banked clinical samples.

Two novel real-time PCR assays were developed for the detection of Rickettsia spp. One assay detects all tested Rickettsia spp.; the other is specific for Rickettsia rickettsii. Evaluation using DNA from human blood and tissue samples showed both assays to be more sensitive than nested PCR assays currently in use at the CDC.

Presence and persistence of Coxiella burnetii in the environments of goat farms associated with a Q fever outbreak.

Q fever is a zoonotic disease caused by inhalation of the bacterium Coxiella burnetii. Ruminant livestock are common reservoirs for C. burnetii, and bacteria present in aerosols derived from the waste of infected animals can infect humans. The significance of infection from material deposited in the environment versus transmission directly from infected animals is not known. In 2011, an outbreak of Q fever cases on farms in Washington and Montana was associated with infected goats. A study was undertaken to investigate the quantity and spatial distribution of C. burnetii in the environment of these goat farms. Soil, vacuum, and sponge samples collected on seven farms epidemiologically linked to the outbreak were tested for the presence of C. burnetii DNA by quantitative PCR. Overall, 70.1% of the samples were positive for C. burnetii. All farms had positive samples, but the quantity of C. burnetii varied widely between samples and between farms. High quantities of C. burnetii DNA were in goat housing/birthing areas, and only small quantities were found in samples collected more than 50 m from these areas. Follow-up sampling at one of the farms 1 year after the outbreak found small quantities of C. burnetii DNA in air samples and large quantities of C. burnetii persisting in soil and vacuum samples. The results suggest that the highest concentrations of environmental C. burnetii are found in goat birthing areas and that contamination of other areas is mostly associated with human movement.

Stability of Coxiella burnetii in stored human blood.

BACKGROUND: Coxiella burnetii, an obligate intracellular organism, is the causative agent of the zoonotic disease Q fever. The seroprevalence rate for Q fever in the United States is 3.1%, suggesting a high number of infections each year. However, fewer than 200 cases of Q fever are reported to the CDC annually. This discrepancy is likely the result of underutilized diagnostics and a high percentage (>50%) of asymptomatic infections. The detection of C. burnetii in patient blood during the first 2 to 3 weeks of infection raises the possibility that the organism could be present in donated human blood. The purpose of this study was to determine if extracellular C. burnetii would be stable in blood under normal storage conditions. STUDY DESIGN AND METHODS: Donated human blood was separated into whole blood, leukoreduced whole blood, red blood cells, and plasma. Each component was spiked with purified, extracellular C. burnetii strain Nine Mile Phase 1, and the viability and infectivity of the organisms were tested weekly. RESULTS: C. burnetii did not decrease in viability or the ability to infect cells after storage in any of the blood products, even after 6 weeks of storage at 1 to 6 °C. CONCLUSIONS: Extracellular C. burnetii can survive and remain infectious in donated blood products.

Molecular typing of Coxiella burnetii (Q fever).

Although we live in the age of genomics and the availability of complete genome sequences of Coxiella burnetii has increased our understanding of the genomic diversity of the agent, it is still somewhat a "query" microorganism. The epidemiology of Q fever is complex due to the worldwide distribution, reservoir and vector diversity, and a lack of studies defining the dynamic interaction between these factors. In addition Coxiella is an agent that could be used as a bioterror weapon. Therefore, typing methods that can discriminate strains and be used to trace back infections to their source are of paramount importance. In this chapter we provide an overview of historical and current typing methods and describe their advantages and limitations. Recently developed techniques such as MLVA and SNP typing have shown promise and improved the discrimination capacity and utility of genotyping methods for molecular epidemiologic studies of this challenging pathogen.

Practice guidelines for the diagnosis and management of patients with Q fever by the Armed Forces Infectious Diseases Society.

This issue in the series Current Topics in Military Tropical Medicine focuses on Q Fever. Q fever is a zoonotic infection caused by the bacterium Coxiella burnetii. Over 150 confirmed cases have occurred among U.S. military personnel deployed to Iraq since 2007. Acute Q fever is underdiagnosed because of a myriad of possible clinical presentations but typically presents as a flu-like illness. The most common chronic manifestation is endocarditis. Most providers are not familiar with the diagnosis, treatment, or appropriate follow-up of this disease. In order to facilitate the care of patients infected with C. burnetii, the Armed Forces Infectious Diseases Society convened a panel of experts in the field to develop practical guidelines for those caring for infected patients. The recommendations and rationale are reviewed in this article.

Q fever, spotted fever group, and typhus group rickettsioses among hospitalized febrile patients in northern Tanzania.

BACKGROUND: The importance of Q fever, spotted fever group rickettsiosis (SFGR), and typhus group rickettsiosis (TGR) as causes of febrile illness in sub-Saharan Africa is unknown; the putative role of Q fever as a human immunodeficiency virus (HIV) coinfection is unclear. METHODS: We identified febrile inpatients in Moshi, Tanzania, from September 2007 through August 2008 and collected acute- and convalescent-phase serum samples. A ≥4-fold increase in immunoglobulin (Ig) G immunfluorescence assay (IFA) titer to Coxiella burnetii phase II antigen defined acute Q fever. A ≥4-fold increase in IgG IFA titer to Rickettsia conorii or Rickettsia typhi antigen defined SFGR and TGR, respectively. RESULTS: Among 870 patients, 483 (55.5%) were tested for acute Q fever, and 450 (51.7%) were tested for acute SFGR and TGR. Results suggested acute Q fever in 24 (5.0%) patients and SFGR and TGR in 36 (8.0%) and 2 (0.5%) patients, respectively. Acute Q fever was associated with hepato- or splenomegaly (odds ratio [OR], 3.1; P = .028), anemia (OR, 3.0; P = .009), leukopenia (OR, 3.9; P = .013), jaundice (OR, 7.1; P = .007), and onset during the dry season (OR, 2.7; P = .021). HIV infection was not associated with acute Q fever (OR, 1.7; P = .231). Acute SFGR was associated with leukopenia (OR, 4.1; P = .003) and with evidence of other zoonoses (OR, 2.2; P = .045). CONCLUSIONS: Despite being common causes of febrile illness in northern Tanzania, Q fever and SFGR are not diagnosed or managed with targeted antimicrobials. C. burnetii does not appear to be an HIV-associated co-infection.

Brill-Zinsser disease in a patient following infection with sylvatic epidemic typhus associated with flying squirrels.

Recrudescent Rickettsia prowazekii infection, also known as Brill-Zinsser disease, can manifest decades after untreated primary infection but is rare in contemporary settings. We report the first known case of Brill-Zinsser disease in a patient originally infected with a zoonotic strain of R. prowazekii acquired from flying squirrels.

Coxiella burnetii infection of a Steller sea lion (Eumetopias jubatus) found in Washington State.

A pregnant sea lion stranded in the State of Washington was found to have placentitis caused by a unique strain of Coxiella burnetii. This is the first description of coxiellosis in a sea lion and suggests that exposure to sea lions may be a risk factor for contracting Q fever.

Practical method for extraction of PCR-quality DNA from environmental soil samples.

Methods for the extraction of PCR-quality DNA from environmental soil samples by using pairs of commercially available kits were evaluated. Coxiella burnetii DNA was detected in spiked soil samples at <1,000 genome equivalents per gram of soil and in 12 (16.4%) of 73 environmental soil samples.

Presence of Coxiella burnetii DNA in the environment of the United States, 2006 to 2008.

Coxiella burnetii is an obligate intracellular bacterium that causes the zoonotic disease Q fever. Because C. burnetii is highly infectious, can survive under a variety of environmental conditions, and has been weaponized in the past, it is classified as a select agent and is considered a potential bioweapon. The agent is known to be present in domestic livestock and in wild animal populations, but the background levels of C. burnetii in the environment have not been reported. To better understand the amount of C. burnetii present in the environment of the United States, more than 1,600 environmental samples were collected from six geographically diverse parts of the United States in the years 2006 to 2008. DNA was purified from these samples, and the presence of C. burnetii DNA was evaluated by quantitative PCR of the IS1111 repetitive element. Overall, 23.8% of the samples were positive for C. burnetii DNA. The prevalence in the different states ranged from 6 to 44%. C. burnetii DNA was detected in locations with livestock and also in locations with primarily human activity (post offices, stores, schools, etc.). This study demonstrates that C. burnetii is fairly common in the environment in the United States, and any analysis of C. burnetii after a suspected intentional release should be interpreted in light of these background levels. It also suggests that human exposure to C. burnetii may be more common than what is suggested by the number of reported cases of Q fever.

Detection of Coxiella burnetii in commercially available raw milk from the United States.

Unpasteurized (raw) milk can be purchased in 39 U.S. states, with direct consumer purchase for human consumption permitted in 29 of those 39 states. Raw milk (n=21; cow, 14; goat, 7) was purchased in 12 states, and Coxiella burnetii, the agent of Q fever, was detected in 9 of 21 (42.9%) samples tested by polymerase chain reaction. Viability of the pathogen was demonstrated by isolation of the agent in tissue culture. The demonstration of viable C. burnetii in commercially available raw milk poses a potential public health risk.

Seroepidemiologic and occupational risk survey for Coxiella burnetii antibodies among US veterinarians.

BACKGROUND: Little is known about the occurrence of Q fever among veterinarians in the United States. In this study, we sought to estimate the prevalence of Coxiella burnetii antibodies among veterinarians and to identify risk factors for exposure. METHODS: We tested serum samples from 508 veterinarians who attended the 143rd American Veterinary Medical Association Annual Convention in 2006. Samples were screened using a Q fever IgG enzyme-linked immunosorbent assay (ELISA). Samples with positive or equivocal results of ELISA were confirmed using phase I and phase II IgG immunofluorescence antibody assays, and end point IgG titers were determined for samples with positive results. RESULTS: Antibodies against C. burnetii were detected in 113 (22.2%) of 508 veterinarians. Risk factors associated with seropositivity included age 46 years, routine contact with ponds, and treatment of cattle, swine, or wildlife. CONCLUSIONS: Veterinarians have a high level of exposure to C. burnetii, the causative organism of Q fever, especially those veterinarians who treat livestock. In this study, risk of C. burnetii seropositivity was also independently associated with contact with ponds. The role of exposure to standing bodies of water in infection is not usually considered and should be investigated in future studies. Additionally, the evidence of past infection with C. burnetii in >20% of veterinarians also highlights the need for use of appropriate personal protective equipment when treating animals that are potentially infected with C. burnetii. Physicians should consider the risk of infection with C. burnetii when treating ill veterinarians and others with potential occupational exposures.

Diversity of Anaplasma phagocytophilum strains, USA.

We analyzed the structure of the expression site encoding the immunoprotective protein MSP2/P44 from multiple Anaplasma phagocytophilum strains in the United States. The sequence of p44ESup1 had diverged in Ap-variant 1 strains infecting ruminants. In contrast, no differences were detected between A. phagocytophilum strains infecting humans and domestic dogs.

Succinate dehydrogenase gene arrangement and expression in Anaplasma phagocytophilum.

DNA sequencing of the region directly downstream of the Anaplasma phagocytophilum (strain MRK) 16S rRNA gene identified homologues of sdhC and sdhD; however, further sequencing by gene walking failed to identify additional sdh gene homologues. The sequence downstream of sdhD identified a partial gene, pep1, predicted to encode a protein >35.3 kDa with 26.3% identity to a hypothetical Ehrlichia canis protein with no known function. The recently completed sequence of the A. phagocytophilum genome confirmed our findings and indicated that the sdhA and sdhB genes are duplicated in a tandem orientation, and located distant from the sdhC and sdhD genes. The expression of the A. phagocytophilum 16S rRNA, sdhC, and sdhD genes was examined by reverse transcriptase PCR which showed that these three genes are expressed as an operon. The pep1 gene was expressed independent of the 16S-sdhCD operon from a promoter between sdhD and pep1. Further analysis of the sdhA and sdhB genes suggested the tandem duplication of the genes in conserved and may be unique to the species A. phagocytophilum. While the conservation of the A. phagocytophilum Sdh proteins, including the residues required for heme- and quinone-binding by SdhC and SdhD, suggests these subunits form an active enzymatic complex, the unusual genomic arrangement and expression pattern of these genes support previous studies (rRNA, ftsZ) indicating that gene rearrangement and operon fragmentation are common in the genomes of Anaplasma and other obligate intracellular bacteria. OMB DISCLAIMER: The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the CDC or the Department of Health and Human Services.

A sustained-release formulation of doxycycline hyclate (Atridox) prevents simultaneous infection of Anaplasma phagocytophilum and Borrelia burgdorferi transmitted by tick bite.

Current prophylaxis for infected tick bites consists of personal protective measures directed towards ticks. This study compared the efficacy of a single oral dose of doxycycline with that of a single injection of sustained-release doxycycline in a model of Lyme borreliosis and Anaplasma phagocytophilum infection. Dosages of doxycycline were equilibrated based on previously determined peak plasma levels in mice [oral, 2.4 microg (ml plasma)(-1); sustained release, 1.9 microg (ml plasma)(-1)] determined 8 h after inoculation. In challenge experiments where five Borrelia burgdorferi-infected and five A. phagocytophilum-infected nymphs were used per mouse, only 20 and 30 % of mice were protected from B. burgdorferi and A. phagocytophilum infection, respectively, using oral doxycycline. In contrast, 100 % of mice receiving sustained-release doxycycline were protected from A. phagocytophilum infection, as indicated by real-time PCR of blood samples, quantitative PCR and culture isolation of spleen samples, and protected against B. burgdorferi infection as demonstrated by culture of ear, heart and bladder. Although 15-40 copies of A. phagocytophilum could be amplified from the spleens of mice treated with sustained-release doxycycline, no viable A. phagocytophilum from these spleens could be cultured in HL-60 cells. In contrast, 7/10 mice receiving oral doxycycline were PCR- and culture-positive for A. phagocytophilum, with copy numbers ranging from 800 to 10 000 within the spleen, as determined by quantitative PCR. Other correlates with A. phagocytophilum infection included a significant difference in spleen mass (mean of 110 mg for sustained-release treatment versus a mean of 230 mg for oral treatment) and the number of splenic lymphoid nodules (mean of 8 for sustained-release treatment versus mean of 12.5 for oral doxycycline) as determined by histopathology. These studies indicate that a single injection of a sustained-release formulation antibiotic may offer a viable prophylactic treatment option for multiple infectious agents in patients presenting with tick bites.