BackgroundThe lone star tick Amblyomma americanum is a common pest and vector of infectious diseases for humans and other mammals in the southern and eastern United States. A Coxiella sp. bacterial endosymbiont was highly prevalent in both laboratory-reared and field-collected A. americanum. The Coxiella sp. was demonstrated in all stages of tick and in greatest densities in nymphs and adult females, while a Rickettsia sp. was less prevalent and in lower densities when present.Methodology/Principal FindingsWe manipulated the numbers of both bacterial species in laboratory-reared A. americanum by injecting engorged nymphs or engorged, mated females with single doses of an antibiotic (rifampin or tetracycline) or buffer alone. Burdens of the bacteria after molting or after oviposition were estimated by quantitative polymerase chain reaction with primers and probes specific for each bacterial species or, as an internal standard, the host tick. Post-molt adult ticks that had been treated with rifampin or tetracycline had lower numbers of the Coxiella sp. and Rickettsia sp. and generally weighed less than ticks that received buffer alone. Similarly, after oviposition, females treated previously with either antibiotic had lower burdens of both bacterial species in comparison to controls. Treatment of engorged females with either antibiotic was associated with prolonged time to oviposition, lower proportions of ticks that hatched, lower proportions of viable larvae among total larvae, and lower numbers of viable larvae per tick. These fitness estimators were associated with reduced numbers of the Coxiella sp. but not the Rickettsia sp.Conclusion/SignificanceThe findings indicate that the Coxiella sp. is a primary endosymbiont, perhaps provisioning the obligately hematophagous parasites with essential nutrients. The results also suggest that antibiotics could be incorporated into an integrated pest management plan for control of these and other tick vectors of disease.

Humans and other animals with Lyme borreliosis produce antibodies to a number of components of the agent Borrelia burgdorferi, but a full accounting of the immunogens during natural infections has not been achieved. Employing a protein array produced in vitro from 1,292 DNA fragments representing ϳ80% of the genome, we compared the antibody reactivities of sera from patients with early or later Lyme borreliosis to the antibody reactivities of sera from controls. Overall, ϳ15% of the open reading frame (ORF) products ( Infectious disease research has advanced rapidly with the accumulation of whole-genome sequences of pathogens and the subsequent use of genome-wide DNA microarrays to study gene expression. Equipped with arrays in different formats, investigators have identified different genes in a variety of pathogens that are more highly expressed in host animals or under in vitro conditions mimicking the in vivo environment. With few exceptions (27), these array studies have been performed with experimental animals, usually rodents, and in laboratory settings. Less is known about the proteins that are expressed during natural infections of humans or other host animals. Detection of a specific antibody during the course of infection is indirect evidence of in vivo expression by the pathogen. But the use of this approach to study large numbers of proteins has been largely limited to one-dimensional and two-dimensional gel electrophoresis of whole cells having an in vitro origin, followed by identification of the more abundant antigens by partial amino acid sequencing of reactive bands or spots and then searches of the databases (22,23,38,44,52,60,66).An alternative to using the pathogen itself as the source of the proteins is to produce recombinant polypeptides based on the deduced open reading frames (ORFs) of the pathogen's genome and to determine whether these polypeptides are antibody targets (11,61). A potential shortcoming of using this approach with cells, such as Escherichia coli or yeast (Saccharomyces cerevisiae) cells, is that some foreign proteins may not be expressed or the quantity may be insufficient. Felgner and coinvestigators increased the success rate for expression and decreased the cost with a high-throughput, cell-free, coupled transcription-translation system (26,29,30). Hundreds to thousands of individual recombinant proteins are printed on chips, which are then used to capture antibodies present in serum from infected individuals and other animals. The amount of captured antibody is quantified using a labeled secondary antibody. Whole-proteome microarrays have been employed in studies of experimental Francisella tularensis infections in laboratory mice (35,57) and of immune responses of humans to immunization with live vaccinia virus (26,29,31). McKevitt et al. (61) and Brinkmann et al. (11) used the enzyme-linked immunosorbent assay (ELISA) format to study the binding of antibodies of experimentally infected rabbits and people with syphilis to a nearly complete representation of the OR...

Laboratory-reared and field-collected Amblyomma americanum ticks were hosts of a Coxiella sp. and a Rickettsia sp. While the Coxiella sp. was detected in 50 of 50 field-collected ticks, the Rickettsia sp. was absent from 32% of ticks. The Coxiella sp. showed evidence of a reduced genome and may be an obligate endosymbiont.

Antigen profiling using comprehensive protein microarrays is a powerful tool for characterizing the humoral immune response to infectious pathogens. Coxiella burnetii is a CDC category B bioterrorist infectious agent with worldwide distribution. In order to assess the antibody repertoire of acute and chronic Q fever patients we have constructed a protein microarray containing 93% of the proteome of Coxiella burnetii, the causative agent of Q fever. Here we report the profile of the IgG and IgM seroreactivity in 25 acute Q fever patients in longitudinal samples. We found that both early and late time points of infection have a very consistent repertoire of IgM and IgG response, with a limited number of proteins undergoing increasing or decreasing seroreactivity. We also probed a large collection of acute and chronic Q fever patient samples and identified serological markers that can differentiate between the two disease states. In this comparative analysis we confirmed the identity of numerous IgG biomarkers of acute infection, identified novel IgG biomarkers for acute and chronic infections, and profiled for the first time the IgM antibody repertoire for both acute and chronic Q fever. Using these results we were able to devise a test that can distinguish acute from chronic Q fever. These results also provide a unique perspective on isotype switch and demonstrate the utility of protein microarrays for simultaneously examining the dynamic humoral immune response against thousands of proteins from a large number of patients. The results presented here identify novel seroreactive antigens for the development of recombinant protein-based diagnostics and subunit vaccines, and provide insight into the development of the antibody response. Molecular & Cellular

Studying tick feeding and digestion, we discovered in a cDNA library from partially fed Amblyomma americanum ticks the first known arthropod homologue of a human cytokine, the pro-inflammatory Macrophage Migration Inhibitory Factor (MIF). The tick origin of the MIF cDNA clone was confirmed by sequencing a genomic fragment that contained the full-length tick MIF gene with two introns. Antiserum to a tick MIF-specific peptide as well as antiserum to complete tick MIF revealed the expression of tick MIF in the salivary gland and midgut tissues of A. americanum ticks. In an in vitro functional assay, recombinant tick MIF inhibited the migration of human macrophages to the same extent that recombinant human MIF did.