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Two Hepatozoon spp are recognized as parasites of domestic dogs in the United States, H. canis and H. americanum. H. canis was first described in India in 1905 and has been documented in many areas of the world, although not definitively identified in North America until recently. H. americanum, causing American canine hepatozoonosis, was first documented in a coyote in 1978 and is now considered an emerging etiologic agent of disease in domestic dogs throughout the United States. The authors review current knowledge of canine hepatozoonosis caused by H. canis and H. americanum and elaborate on more recent research findings.

Genital herpes is one of the most common sexually transmitted infections, affecting 1 in 6 people in the United States. Women are twice as likely to be infected as men and infections in women of reproductive age carry the additional risk of vertical transmission to the neonate at the time of delivery. Neonatal herpes infections can be devastating with up to 50% mortality for disseminated herpes simplex virus (HSV) infections in the newborn. Rates of transmission are affected by the viral type of HSV infection and whether the infection around delivery is primary or recurrent. Current management approaches decrease rates of active lesions at the time of delivery and thereby cesarean deliveries, but have not been shown to decrease the incidence of neonatal herpes infections. More research is needed to better elucidate the risk factors for transmission to the neonate and to improve our current management methodology to further decrease vertical transmission. In this review, we will discuss management of antenatal and peripartum herpes infections, considerations for mode of delivery, and the course of neonatal HSV infections., (Copyright © 2011 by Lippincott Williams & Wilkins)

Women at risk for Lynch Syndrome/HNPCC have an increased lifetime risk of endometrial and ovarian cancer. This study investigates the cost-effectiveness of prophylactic surgery versus surveillance in women with Lynch Syndrome. A decision analytic model was designed incorporating key clinical decisions and existing probabilities, costs, and outcomes from the literature. Clinical forum where risk-reducing surgery and surveillance were considered. A theoretical population of women with Lynch Syndrome at age 30 was used for the analysis. A decision analytic model was designed comparing the health outcomes of prophylactic hysterectomy with bilateral salpingo-oophorectomy at age 30 versus annual gynecologic screening versus annual gynecologic exam. The literature was searched for probabilities of different health outcomes, results of screening modalities, and costs of cancer diagnosis and treatment. Cost-effectiveness expressed in dollars per discounted life-years. Risk-reducing surgery is the least expensive option, costing $23,422 per patient for 25.71 quality-adjusted life-years (QALYs). Annual screening costs $68,392 for 25.17 QALYs; and annual examination without screening costs $100,484 for 24.60 QALYs. Further, because risk-reducing surgery leads to both the lowest costs and the highest number of QALYs, it is a dominant strategy. Risk-reducing surgery is the most cost-effective option from a societal healthcare cost perspective.

Short-term consumption of a high-fat diet impairs exercise capacity in both rats and humans, and increases expression of the mitochondrial uncoupling protein, UCP3, in rodent cardiac and skeletal muscle via activation of the transcription factor, peroxisome proliferator-activated receptor α (PPARα). Unlike long-chain fatty acids however, medium-chain fatty acids do not activate PPARα and do not increase muscle UCP3 expression. We therefore investigated exercise performance and cardiac mitochondrial function in rats fed a chow diet (7.5% kcal from fat), a long-chain triglyceride (LCT) rich diet (46% kcal from LCTs) or a medium-chain triglyceride (MCT) rich diet (46% kcal from MCTs). Rats fed the LCT-rich diet for 15 days ran 55% less far than they did at baseline, whereas rats fed the chow or MCT-rich diets neither improved nor worsened in their exercise capacities. Moreover, consumption of an LCT-rich diet increased cardiac UCP3 expression by 35% and decreased oxidative phosphorylation efficiency, whereas consumption of the MCT-rich diet altered neither UCP3 expression nor oxidative phosphorylation efficiency. Our results suggest that the negative effects of short-term high-fat feeding on exercise performance are predominantly mediated by long-chain rather than medium-chain fatty acids, possibly via PPARα-dependent upregulation of UCP3.

Novel Hepatozoon spp. sequences collected from previously unrecognized vertebrate hosts in North America were compared with documented Hepatozoon 18S rRNA sequences in an effort to examine phylogenetic relationships between the different Hepatozoon organisms found cycling in nature. An approximately 500-base pair fragment of 18S rDNA common to Hepatozoon spp. and some other apicomplexans was amplified and sequenced from the tissues or blood of 16 vertebrate host species from the southern United States, including 1 opossum (Didelphis virginiana), 2 bobcats (Lynx rufus), 1 domestic cat (Felis catus), 3 coyotes (Canis latrans), 1 gray fox (Urocyon cinereoargenteus), 4 raccoons (Procyon lotor), 1 pet boa constrictor (Boa constrictor imperator), 1 swamp rabbit (Sylvilagus aquaticus), 1 cottontail rabbit (Sylvilagus floridanus), 4 woodrats (Neotoma fuscipes and Neotoma micropus), 3 white-footed mice (Peromyscus leucopus), 8 cotton rats (Sigmodon hispidus), 1 cotton mouse (Peromyscus gossypinus), 1 eastern grey squirrel (Sciurus carolinensis), and 1 woodchuck (Marmota monax). Phylogenetic analyses and comparison with sequences in the existing database revealed distinct groups of Hepatozoon spp., with clusters formed by sequences obtained from scavengers and carnivores (opossum, raccoons, canids, and felids) and those obtained from rodents. Surprisingly, Hepatozoon spp. sequences from wild rabbits were most closely related to sequences obtained from carnivores (97.2% identical), and the sequence from the boa constrictor was most closely related to the rodent cluster (97.4% identical). These data are consistent with recent work identifying prey-predator transmission cycles in Hepatozoon spp. and suggest this pattern may be more common than previously recognized.

In this study, the infection dynamics of Ehrlichia ewingii, causative agent of granulocytotropic ehrlichiosis in dogs and humans, was examined in experimentally infected dogs by using a combination of physical examination, hematologic and biochemical analyses, and molecular and serologic assays. For the experimental trials, blood from an E. ewingii-infected dog was inoculated intravenously into two naïve dogs and two dogs with prior experimental exposure to E. ewingii (both were negative for E. ewingii DNA by polymerase chain reaction (PCR) assay, but seropositive from initial infection 8 and 10 months prior to challenge). A negative control dog was inoculated with blood from a negative dog. The two primary infection dogs were positive for E. ewingii DNA on DPI 4, remained consistently positive until DPI 60, and were intermittently positive until the end of the study (DPI 144). The two primary infection dogs developed antibodies reactive to E. ewingii by DPI 28 and remained seropositive for the duration of the study. Primary infected dogs had intermittent fever, thrombocytopenia, and leukopenia and some dogs were hyperphosphatemic and/or had elevated ALP levels. The two challenge dogs were positive for E. ewingii DNA on DPI 4 and 18, which was similar to the primary infection dogs, but the duration of E. ewingii DNA detection was shorter. Also, the two challenged dogs did not develop pyrexia or show any hematologic or biochemical abnormalities. E. ewingii was successfully transmitted between dogs by Amblyomma americanum, but not Rhipicephalus sanguineus. This study provides data on the infection dynamics of E. ewingii in dogs during primary and challenge infections and suggests that prior exposure may lessen clinical disease during subsequent infections., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Objective: To investigate the cost-effectiveness of elective induction of labor at 41 weeks in nulliparous women., Study Design: A decision analytic model comparing induction of labor at 41 weeks vs expectant management with antenatal testing until 42 weeks in nulliparas was designed. Baseline assumptions were derived from the literature as well as from analysis of the National Birth Cohort dataset and included an intrauterine fetal demise rate of 0.12% in the 41st week and a cesarean rate of 27% in women induced at 41 weeks. One-way and multiway sensitivity analyses were conducted to examine the robustness of the findings., Results: Compared with expectant management, induction of labor is cost-effective with an incremental cost of $10,945 per quality-adjusted life year gained. Induction of labor at 41 weeks also resulted in a lower rate of adverse obstetric outcomes, including neonatal demise, shoulder dystocia, meconium aspiration syndrome, and severe perineal lacerations., Conclusion: Elective induction of labor at 41 weeks is cost-effective and improves outcomes., (Copyright © 2011 Mosby, Inc. All rights reserved.)

There is no labeled treatment for dogs with American canine hepatozoonosis (ACH), but the drug therapies discussed in this article, although not rapidly curative, may be successful in alleviating acute clinical signs, prolonging life, reducing the number of clinical relapses, and enhancing quality of life. This article also describes a pilot trial conducted to assess the efficacy of a novel treatment approach with ponazuril as a stand-alone parasiticide administered for 4 weeks without follow-up decoquinate treatment. Although extended ponazuril treatment in combination with NSAID administration did ameliorate acute clinical signs associated with ACH, the parasite was not completely cleared with this treatment protocol alone. Long-term decoquinate therapy remains a critical component of successful treatment of ACH.

Sensitivity to temozolomide is restricted to a subset of glioblastoma patients, with the major determinant of resistance being a lack of promoter methylation of the gene encoding the repair protein DNA methyltransferase MGMT, although other mechanisms are thought to be active. There are, however, limited preclinical data in model systems derived from pediatric glioma patients. We screened a series of cell lines for temozolomide efficacy in vitro, and investigated the differential mechanisms of resistance involved. In the majority of cell lines, a lack of MGMT promoter methylation and subsequent protein overexpression were linked to temozolomide resistance. An exception was the pediatric glioblastoma line KNS42. Expression profiling data revealed a coordinated upregulation of HOX gene expression in resistant lines, especially KNS42, which was reversed by phosphoinositide 3-kinase pathway inhibition. High levels of HOXA9/HOXA10 gene expression were associated with a shorter survival in pediatric high-grade glioma patient samples. Combination treatment in vitro of pathway inhibition and temozolomide resulted in a highly synergistic interaction in KNS42 cells. The resistance gene signature further included contiguous genes within the 12q13-q14 amplicon, including the Akt enhancer PIKE, significantly overexpressed in the KNS42 line. These cells were also highly enriched for CD133 and other stem cell markers. We have thus shown an in vitro link between phosphoinositide 3-kinase-mediated HOXA9/HOXA10 expression, and a drug-resistant, progenitor cell phenotype in MGMT-independent pediatric glioblastoma., (Copyright © 2010 AACR.)

In the time since canine ehrlichiosis due to Ehrlichia canis was first described in 1935 and first recognized in the United States in 1962, many key advances have been made in our understanding of the diversity of the rickettsial organisms responsible for ehrlichiosis and anaplasmosis in dogs and, occasionally, cats, the vectors capable of transmitting these agents, and the role these organisms play as both important veterinary pathogens and zoonotic disease agents. Despite considerable progress in the field, much remains to be learned regarding mechanisms contributing to pathogenesis, effective treatment modalities, and prevention strategies that best protect pet health. This article highlights current understanding of the transmission, diagnosis, and management of ehrlichiosis and anaplasmosis in dogs and cats., (Copyright © 2010 Elsevier Inc. All rights reserved.)

We used PCR and a novel serologic assay to determine infection and exposure rates to Ehrlichia ewingii in dogs from an area of northeast Oklahoma and northwest Arkansas where Amblyomma americanum ticks are abundant. Of 143 dogs assayed, 13 (9.1%) harbored E. ewingii by PCR and 64 (44.8%) had antibodies to E. ewingii detected using a peptide-based microtiter plate ELISA. Dogs were more likely (P=0.001) to be positive by PCR if sampled in August (30.8%) but no association was found between seropositive status and month of collection of sample (P>0.05). Additional testing revealed PCR evidence of Ehrlichia chaffeensis (4/143; 2.8%) and Anaplasma platys (5/143; 3.5%) as well as antibodies reactive to E. chaffeensis (25/143; 17.5%), Ehrlichia canis (2/143; 1.4%), and Anaplasma spp. (8/143; 5.6%). Testing of another 200 dogs from the area revealed additional PCR and/or serologic evidence of E. ewingii, E. canis, E. chaffeensis, and A. platys. None of the 343 dogs evaluated had evidence of Borrelia burgdorferi exposure. These data support the interpretation that E. ewingii may be the primary agent of canine ehrlichiosis in this region, and suggest that diagnostic evaluation of dogs suspected to have a tick-borne disease should include assays targeting this organism.

Objective: To determine the optimal gestational age of delivery for women with placenta previa by accounting for both neonatal and maternal outcomes., Study Design: A decision-analytic model was designed comparing total maternal and neonatal quality-adjusted life years for delivery of women with previa at gestational ages from 34 to 38 weeks. At each week, we allowed for four different delivery strategies: (1) immediate delivery, without amniocentesis or steroids; (2) delivery 48 hours after steroid administration (without amniocentesis); (3) amniocentesis with delivery if fetal lung maturity (FLM) positive or retesting in one week if FLM negative; (4) amniocentesis with delivery if FLM testing is positive or administration of steroids if FLM negative., Results: Delivery at 36 weeks, 48 hours after steroids, for women with previa optimizes maternal and neonatal outcomes. In sensitivity analyses, these results were robust to a wide range of variation in input assumptions. If it is assumed that steroids offer no neonatal benefit at this gestational age, outright delivery at 36 weeks' gestation is the best strategy., Conclusion: Steroid administration at 35 weeks and 5 days followed by delivery at 36 weeks for women with placenta previa optimizes maternal and neonatal outcomes.

The majority of emerging diseases in humans have been linked to zoonotic pathogens originating in domestic animals or wildlife. This is a public health concern because zoonotic infections affect several aspects of the society. The complex interactions among pathogen, host and environment also pose challenges in estimating the true burden of those infections. However, the recent development of new molecular diagnostic tools has allowed for better diagnosis of zoonotic infections. This review focuses on 3 emerging zoonoses, namely toxocariasis, bovine tuberculosis and southern tick-associated rash illness, and demonstrates that these infections may be more prevalent in the southern United States than previously recognized. This review places special emphasis on the recent epidemiologic trends, intra/interspecies transmission and clinical features of each of these zoonoses. In addition, treatment and prevention for each zoonotic pathogen are discussed. Clinicians working in the southern United States should be aware of the presence of those zoonotic infections.

Established and emerging tickborne infections are significant causes of human illness in the southern United States. Rocky Mountain spotted fever, human monocytic ehrlichiosis, human ewingii ehrlichiosis and tularemia are known pathogens in this geographic distribution. Rickettsia parkeri and novel ehrlichioses are more recently described tickborne infections reviewed in this article. An understanding of the tick vectors, pathogenesis, epidemiology, clinical presentation, diagnosis and treatment is useful for the clinician treating patients potentially infected with any of these pathogens. Prevention measures, the optimal method for removing an attached tick and current and future vaccine development conclude this review.

Amblyomma americanum was confirmed as a competent vector in the transmission of Cytauxzoon felis to domestic cats. Infection with C. felis was produced and replicated in four domestic felines by the bite of A. americanum adults that were acquisition fed as nymphs on a domestic cat that survived cytauxzoonosis. Numerous attempts to transmit C. felis with Dermacentor variabilis at the same time were not successful. All cats upon which infected A. americanum were transmission fed exhibited disease typical of cytauxzoonosis, and the eitiologic agent's presence was confirmed. Clinical signs including fever, inappetence, depression, and lethargy were observed beginning 13 d postinfestation. Pale mucus membranes, splenomegaly, icterus, and dyspnea were also observed during the course of the disease. Rectal temperatures of the C. felis-infected principal cats fluctuated from high to subnormal before returning to the normal range. Clinical signs of cytauxzoonsis improved by 24 d postinfestation in all but one cat, with survivors remaining parasitemic and subclinically infected with C. felis. Unengorged A. americanum and D. variabilis were collected from wild habitats to determine the minimum infection rate of C. felis in ticks from an enzootic area. Infection of C. felis was found only in wild-collected A. americanum. The minimum infection rate of C. felis in A. americanum was 0.5% (one of 178) in males, 0.8% (three of 393) in nymphs, and 1.5% (three of 197) in females. We found no wild-collected D. variabilis infected with C. felis. Our results confirm that A. americanum is a primary vector of C. felis.

Purpose: As genome-scale technologies begin to unravel the complexity of the equivalent tumors in adults, we can attempt detailed characterization of high-grade gliomas in children, that have until recently been lacking. Toward this end, we sought to validate and extend investigations of the differences between pediatric and adult tumors., Experimental Design: We carried out copy number profiling by array comparative genomic hybridization using a 32K bacterial artificial chromosome platform on 63 formalin-fixed paraffin-embedded cases of high-grade glioma arising in children and young people (<23 years)., Results: The genomic profiles of these tumors could be subclassified into four categories: those with stable genomes, which were associated with a better prognosis; those with aneuploid and those with highly rearranged genomes; and those with an amplifier genotype, which had a significantly worse clinical outcome. Independent of this was a clear segregation of cases with 1q gain (more common in children) from those with concurrent 7 gain/10q loss (a defining feature of adults). Detailed mapping of all the amplification and deletion events revealed numerous low-frequency amplifications, including IGF1R, PDGFRB, PIK3CA, CDK6, CCND1, and CCNE1, and novel homozygous deletions encompassing unknown genes, including those at 5q35, 10q25, and 22q13. Despite this, aberrations targeting the "core signaling pathways" in adult glioblastomas are significantly underrepresented in the pediatric setting., Conclusions: These data highlight that although there are overlaps in the genomic events driving gliomagenesis of all ages, the pediatric disease harbors a distinct spectrum of copy number aberrations compared with adults.

Objective: To evaluate the neonatal outcomes of infants delivered to mothers with early-onset preeclampsia., Study Design: This is a retrospective cohort of 1709 infants delivered at 24 0/7 to 29 6/7 weeks gestation was examined. Neonatal outcomes of 235 infants delivered prematurely because of preeclampsia were compared with 1474 infants delivered preterm because of other etiologies. Primary outcomes examined included: small for gestational age (SGA), respiratory distress syndrome (RDS), and neonatal death (NND). Multivariable logistic regression was used to analyze the association between preeclampsia and the neonatal outcomes, controlling for potential confounders., Results: Infants of women with preeclampsia were more likely to be SGA (17.8% vs. 5.6%, AOR 3.9, CI 2.5-6.2) and have RDS (70.6% vs. 60.7%, AOR 1.5, 95% CI 1.1-2.2); however, they were less likely to suffer a NND (11.1% vs. 18.1%, AOR 0.6, 95% CI 0.4-0.9)., Conclusion: Compared with neonates delivered prematurely because of other etiologies, neonates born to preeclamptic mothers were more likely to be SGA and have RDS, but had a decrease in mortality. This may be a reflection of the differences in the underlying pathophysiology behind indicated preterm birth due to preeclampsia.

Dogs and people are exposed to and susceptible to infection by many of the same tick-borne bacterial pathogens in the order Rickettsiales, including Anaplasma phagocytophilum, Ehrlichia canis, E. chaffeensis, E. ewingii, Rickettsia rickettsii, R. conorii, and other spotted fever group rickettsiae. Recent findings include descriptions of novel Ehrlichia and Rickettsia species, recognition of the occurrence and clinical significance of co-infection, and increasing awareness of Rhipicephalus sanguineus-associated diseases. Newer molecular assays are available, although renewed efforts to encourage their use are needed. This review highlights the ecology and epidemiology of these diseases, and proposes avenues for future investigation., (Published by Elsevier Ltd.)

Borrelia burgdorferi sensu stricto is the only established etiologic agent of Lyme borreliosis in dogs and in humans in North America. Lyme borreliosis differs in dogs and humans in terms of clinical outcome following infection, diagnostic approaches, prevention strategies and treatment recommendations. Nonetheless, serologic evidence of exposure of dogs to B. burgdorferi agrees with the geographical distribution of autochthonous transmission of the agent of Lyme borreliosis, and continued monitoring of exposure rates in dogs might allow early recognition of geographic expansion of endemic areas as well as identify hyperendemic areas where both humans and dogs are at increased risk of infection., (Copyright (c) 2010. Published by Elsevier Ltd.)

The lone star tick, Amblyomma americanum (Acari: Ixodidae), is commonly reported from people and animals throughout the eastern U.S. and is associated with transmission of a number of emerging diseases. To better define the microbial communities within lone star ticks, 16S rRNA gene based analysis using bacteria-wide primers, followed by sequencing of individual clones (n = 449) was used to identify the most common bacterial operational taxonomic units (OTUs) present within colony-reared and wild A. americanum. The colony-reared ticks contained primarily sequence affiliated with members of the genus Coxiella (89%; 81/91), common endosymbionts of ticks, and Brevibacterium (11%; 10/91). Similarly, analysis of clones from unfed wild lone star ticks revealed that 96.7% (89/92) of all the OTUs identified were affiliated with Coxiella-like endosymbionts, as compared with only 5.1-11.7% (5/98-9/77) of those identified from wild lone star ticks after feeding. In contrast, the proportion of OTUs identified as Rickettsia sp. in wild-caught ticks increased from 2.2% (2/92) before feeding to as high as 46.8% (36/77) after feeding, and all Rickettsia spp. sequences recovered were most similar to those described from the spotted fever group Rickettsia, specifically R. amblyommii and R. massiliae. Additional characterization of the Rickettsiales tick community by polymerase chain reaction, cloning, and sequencing of 17 kDa and gltA genes confirmed these initial findings and suggested that novel Rickettsia spp. are likely present in these ticks. These data provide insight into the overall, as well as the rickettsial community of wild lone star ticks and may ultimately aid in identification of novel pathogens transmitted by A. americanum.

Objective: To estimate the costs of expanding Emergency Medicaid coverage to include postpartum contraception., Methods: A decision-analytic model was developed using three perspectives: the hospital, state Medicaid programs, and society. Our primary outcome was future reproductive health care costs due to pregnancy in the next 5 years. A Markov structure was use to analyze the probability of pregnancy over a 5-year time period. Model inputs were retrieved from the existing literature and local hospital and Medicaid data related to reimbursements. One-way and multiway sensitivity analyses were conducted. A Monte Carlo simulation was performed to incorporate uncertainty from all of the model inputs simultaneously., Results: Over a 5-year period, provision of contraception would save society $17,792 per woman in future pregnancy costs and incur a loss of $367 for hospitals. In states in which 49% of immigrants remain in the area for 5 years, such a program would save state Medicaid $108 per woman., Conclusion: Under federal regulations, new immigrants are restricted to acute, hospital-based care only. Failure to provide the option of contraception postpartum results in increased costs for society and states with long-term immigrants.

Objective: We sought to investigate the cost-effectiveness of prenatal screening for spinal muscular atrophy (SMA)., Study Design: A decision analytic model was created to compare a policy of universal SMA screening to that of no screening. The primary outcome was incremental cost per maternal quality-adjusted life year. Probabilities, costs, and outcomes were estimated through literature review. Univariate and multivariate sensitivity analyses were performed to test the robustness of our model to changes in baseline assumptions., Results: Universal screening for SMA is not cost-effective at $4.9 million per quality-adjusted life year. In all, 12,500 women need to be screened to prevent 1 case of SMA, at a cost of $5.0 million per case averted. Our results were most sensitive to the baseline prevalence of disease., Conclusion: Universal prenatal screening for SMA is not cost-effective. For populations at high risk, such as those with a family history, SMA testing may be a cost-effective strategy., (Copyright 2010 Mosby, Inc. All rights reserved.)

To determine the national, regional, and age-related prevalence of intestinal parasites in dogs presenting to veterinarians in the United States, we reviewed the results of examination via zinc sulfate centrifugal flotation of 1,199,293 canine fecal samples submitted to Antech Diagnostics in 2006. The most commonly identified intestinal parasites were ascarids (2.2%), hookworms (2.5%), whipworms (1.2%), Giardia (4.0%), and Cystoisospora (4.4%). With the exception of whipworms, intestinal parasites were more commonly identified in dogs less than 6 months of age (29.6% positive) as compared to those greater than 1 year of age (6.1% positive) although infections with each parasite considered were identified in all age classes of dogs. Hookworm eggs were most commonly identified in fecal samples submitted from dogs from the South (4.0% positive), whereas ascarid eggs and Giardia cysts were most commonly seen in samples from dogs from the West (2.8% and 6.3% positive, respectively). When compared to previous data from shelter dogs, the prevalence of intestinal helminths, particularly ascarids and hookworms, was greatly suppressed in pet dogs in the southern United States (90-91% reduction) and much less so in dogs in the West (52-78% reduction), perhaps due in part to the routine year-round use of monthly anthelmintics effective at controlling both heartworm infection and intestinal helminths in dogs in the South. Taken together these data indicate that intestinal parasites remain a common, important finding in dogs presenting to veterinary practices although in most of the country infection rates in pet dogs appear to be greatly reduced from the level reported from dogs in animal shelters.

Southern tick-associated rash illness is a Lyme-like syndrome that occurs in the southern states. Borrelia lonestari, which has been suggested as a possible causative agent of southern tick-associated rash illness, naturally infects white-tailed deer (WTD; Odocoileus virginianus) and is transmitted by the lone star tick (Amblyomma americanum). To better understand the prevalence and distribution of Borrelia exposure among WTD, we tested WTD from 21 eastern states for antibodies reactive to B. lonestari using an indirect immunofluorescent antibody assay and Borrelia burgdorferi using the IDEXX SNAP 4Dx test. A total of 107/714 (15%) had antibodies reactive to B. lonestari, and prevalence of antibodies was higher in deer from southern states (17.5%) than in deer from northern states (9.2%). Using the SNAP 4DX test, we found that 73/723 (10%) were positive for B. burgdorferi, and significantly more northern deer (23.9%) were positive compared with southern deer (3.8%). Our data demonstrate that WTD are exposed to both Borrelia species, but antibody prevalence for exposure to the two species differs regionally and distributions correlate with the presence of Ixodes scapularis and A. americanum ticks.

Background: The range of American canine hepatozoonosis (ACH) is expanding from the southern USA northward. Transmission of Hepatozoon americanum occurs by ingestion of infected Gulf Coast ticks, Amblyomma maculatum. The source of the protozoan for the tick remains undetermined; infected dogs are unusual hosts for the tick., Objective: Compare possible sources of infection by field investigations of 2 multiple-dog outbreaks of ACH., Animals: Twenty-eight privately owned dogs (Canis familiaris), 1 coyote (Canis latrans), 31 wild-trapped cotton rats (Sigmodon hispidus), 24 wild-trapped field mice (Peromyscus leucopus), and 9 wild-caught rabbits (Sylvilagus spp.) from sites in eastern Oklahoma were monitored for hepatozoonosis. Six laboratory-raised cotton rats (S. hispidus), 6 Sprague-Dawley rats (Rattus norvegicus), 6 C57BL/6J-Lystbg-J/J mice (Mus musculus), 6 outbred white mice (M. musculus), 6 New Zealand white rabbits (Oryctolagus cuniculus), and 2 dogs were acquired through commercial vendors for experimental transmission trials of H. americanum., Methods: Four of 15 dogs in a rural neighborhood and 5/12 hunting Beagles were confirmed to be infected by blood smear examination, muscle biopsy, and polymerase chain reaction assay of the 18S rRNA gene of Hepatozoon species. Histories and tick host preferences led to field collections of common prey of canids and experimental transmission trials of H. americanum to selected prey (M. musculus, S. hispidus, R. norvegicus, and O. cuniculus)., Results: Dogs with ready access to prey (4/15 dogs) or that were fed prey retrieved from hunts (5/12 hunting Beagles) became infected, providing evidence that predation is an important epidemiologic component of ACH infection. Experimental transmission studies identified a quiescent, infectious stage (cystozoite) of the parasite that provides an alternate mode of transmission to canids through predation, demonstrating that cotton rats, mice, and rabbits but not brown rats may act as paratenic hosts of H. americanum., Conclusions and Clinical Importance: Predation of prey harboring infected A. maculatum or containing cystozoites of H. americanum in their tissues provide 2 modes of transmission of ACH to dogs, putting unconfined dogs at increased risk of infection in endemic areas.

Inflammatory lesions containing parasitic cystozoites developed in multiple organs and tissues of laboratory-raised Oryctolagus cuniculus that were administered approximately 100 sporulated oocysts of Hepatozoon americanum (Oklahoma isolate, GenBank accession AF176836) orally. The predominantly granulomatous inflammatory lesions were detected histologically 8 weeks after exposure to oocysts. Cystozoites, recognized by cresent-shaped, uninucleated bodies surrounded by an accumulation of globular, PAS-positive polysaccharide material, were evident within macrophages as monozoic and dizoic cysts. Neither meronts nor gamonts were detected in any of the laboratory-raised lagomorphs during the 24-week observation period. Nested PCR assay of rabbit tissues for a 488 bp fragment of the 18S rRNA Hepatozoon spp. gene was positive at 8 and 24 weeks post-exposure. The sequence was 97.1% similar with sequence from the H. americanum carrier used to infect ticks. A Hepatozoon-free dog fed tissues from the 24-week post-exposure rabbit principal developed American canine hepatozoonosis. Gamonts were first detected 5 weeks after the dog ingested the rabbit tissues containing cystozoites. PCR assay of blood from the dog was positive for the Hepatozoon spp. gene fragment. Sequencing confirmed that the parasite in the dog was H. americanum.

Four white-tailed deer were inoculated with either the Ap-V1 or NY-18 strain of Anaplasma phagocytophilum. Ixodes scapularis nymphs were then allowed to acquistion feed on the inoculated deer and molt to adults. Only an Ap-V1 infected deer was infected persistently and able to infect nymphal Ixodes scapularis. Molted adult ticks maintained Ap-V1 infection as demonstrated by PCR and microscopy. We report, for the first time, a morphologic description of A. phagocytophilum in I. scapularis.

Purpose: The epidermal growth factor receptor (EGFR) is amplified and overexpressed in adult glioblastoma, with response to targeted inhibition dependent on the underlying biology of the disease. EGFR has thus far been considered to play a less important role in pediatric glioma, although extensive data are lacking. We have sought to clarify the role of EGFR in pediatric high-grade glioma (HGG)., Experimental Design: We retrospectively studied a total of 90 archival pediatric HGG specimens for EGFR protein overexpression, gene amplification, and mutation and assessed the in vitro sensitivity of pediatric glioma cell line models to the small-molecule EGFR inhibitor erlotinib., Results: Amplification was detected in 11% of cases, with corresponding overexpression of the receptor. No kinase or extracellular domain mutations were observed; however, 6 of 35 (17%) cases harbored the EGFRvIII deletion, including two anaplastic oligodendrogliomas and a gliosarcoma overexpressing EGFRvIII in the absence of gene amplification and coexpressing platelet-derived growth factor receptor alpha. Pediatric glioblastoma cells transduced with wild-type or deletion mutant EGFRvIII were not rendered more sensitive to erlotinib despite expressing wild-type PTEN. Phosphorylated receptor tyrosine kinase profiling showed a specific activation of platelet-derived growth factor receptor alpha/beta in EGFRvIII-transduced pediatric glioblastoma cells, and targeted coinhibition with erlotinib and imatinib leads to enhanced efficacy in this model., Conclusions: These data identify an elevated frequency of EGFR gene amplification and EGFRvIII mutation in pediatric HGG than previously recognized and show the likely necessity of targeting multiple genetic alterations in the tumors of these children.

Objective: To determine the prevalence of Babesia gibsoni infection in dogs that were confiscated from dogfighting operations., Design: Cross-sectional study., Animals: 157 pit bull-type dogs that were confiscated as part of dogfighting prosecution cases in Iowa, Michigan, Mississippi, Ohio, Pennsylvania, Virginia, and Washington and 218 randomly selected animal shelter dogs with no known history of dogfighting., Procedures: Blood samples collected from confiscated dogs were tested for infection with B gibsoni by use of a nested PCR assay. Samples that yielded positive results underwent DNA sequencing to confirm infection with B gibsoni. Control blood samples collected from 218 randomly selected dogs in animal shelters (ie, dogs that had no known involvement in dogfighting events) were also analyzed., Results: Results of nested PCR assays indicated that 53 of 157 (33.8%) confiscated dogs were infected with B gibsoni; 1 (0.6%) dog was infected with the canine small Babesia 'Spanish isolate' (also known as Theileria annae). To the authors' knowledge, this is the first report of infection with this small Babesia 'Spanish isolate' in a North American dog. Dogs with scars (indicative of fighting) on the face, head, and forelimbs were 5.5 times as likely to be infected with B gibsoni as were dogs without scars. Of the control dogs, 1 (0.5%) pit bull-type dog was infected with B gibsoni., Conclusions and Clinical Relevance: Results indicated that B gibsoni is a common parasite of dogs confiscated from dogfighting operations and suggested that dogs with a history of fighting should be evaluated for infection with B gibsoni.

Cytauxzoon felis was transmitted to a domestic cat by Amblyomma americanum. The infection was produced by the bite of A. americanum adults that were acquisition fed as nymphs on a domestic cat that naturally survived infection of C. felis. Fever, inappetence, depression, and lethargy were first noted 11 days post-infestation (dpi). Pale mucus membranes, splenomegaly, icterus, and dyspnea were also observed during the course of the disease. The body temperature of the experimentally infected C. felis cat was subnormal from 16 dpi until 24 dpi when it returned to within normal limits. All clinical signs of cytauxzoonsis began to resolve by 23 dpi when the cat became subclinically infected with C. felis. The cat developed a marked, regenerative anemia beginning by 13 dpi and reached a nadir at 20 dpi before recovering. A moderate neutrophilia and marked lymphocytosis also developed between 18 and 26 dpi. Schizonts of C. felis were observed in spleen aspirates of the infected cat at 15 dpi. DNA of C. felis was amplified by real-time PCR starting 17 dpi and piroplasms of C. felis were first noted by light microscopy 18 dpi. Dermacentor variabilis, Ixodes scapularis, and Rhipicephalus sanguineus were also tested in a similar manner at the same time but did not transmit C. felis. Prior to the present study, only D. variabilis had been shown experimentally to transmit infection of C. felis. This is the first report of C. felis being transmitted by A. americanum. The transmission of C. felis infection from one domestic cat to another indicates that domestic cats subclinically infected with C. felis may be a reservoir of infection for naive domestic cats.

Canine hepatozoonosis is caused by Hepatozoon canis and Hepatozoon americanum, apicomplexan parasites transmitted to dogs by ingestion of infectious stages. Although the two agents are phylogenetically related, specific aspects, including characteristics of clinical disease and the natural history of the parasites themselves, differ between the two species. Until recently, H. canis infections had not been clearly documented in North America, and autochthonous infection with H. americanum has yet to be reported outside of the southern United States. However, recent reports demonstrate H. canis is present in areas of North America where its vector tick, Rhipicephalus sanguineus, has long been endemic, and that the range of H. americanum is likely expanding along with that of its vector tick, Amblyomma maculatum; co-infections with the two organisms have also been identified. Significant intraspecific variation has been reported in the 18S rRNA gene sequence of both Hepatozoon spp.-infecting dogs, suggesting that each species may represent a complex of related genogroups rather than well-defined species. Transmission of H. americanum to dogs via ingestion of cystozoites in muscle of infected vertebrates was recently demonstrated, supporting the concept of predation as a means of natural transmission. Although several exciting advances have occurred in recent years, much remains to be learned about patterns of infection and the nature of clinical disease caused by the agents of canine hepatozoonosis in North America.

We evaluated a comprehensive national database that documents canine infection with, or exposure to, four vector-borne disease agents, Dirofilaria immitis, Borrelia burgdorferi, Ehrlichia canis, and Anaplasma phagocytophilum in order to assess geographic trends in rates of positive tests. While the percent positive test results varied by agent in different regions of the United States, with D. immitis antigen and antibodies to E. canis more commonly identified in dogs from the South (3.9% and 1.3%, respectively), and antibody to B. burgdorferi and A. phagocytophilum found more frequently in dogs from the upper Midwest and Northeast (4.0-6.7% and 5.5-11.6%, respectively), evidence of at least one agent was found in dogs from every state considered. Furthermore, each organism also appeared to occur in endemic foci within larger areas of relatively low prevalence. Relocation of infected or previously exposed dogs from endemic regions likely accounts for some of the unexpected geographic distribution seen, although local transmission in previously under-recognized areas of endemicity could also be occurring. Although data were only available from the 48 contiguous states (Alaska and Hawaii were not included), taken together, our results suggest that these disease agents may be present over a wider geographic area, and thus pose greater animal and public health risks, than is currently recognized. Dogs can serve as sentinels to identify the presence of vector-borne disease agents of both veterinary and public health significance.

Background: Induction of labor is on the rise in the U.S., increasing from 9.5 percent in 1990 to 22.1 percent in 2004. Although, it is not entirely clear what proportion of these inductions are elective (i.e. without a medical indication), the overall rate of induction of labor is rising faster than the rate of pregnancy complications that would lead to a medically indicated induction. However, the maternal and neonatal effects of induction of labor are unclear. Many studies compare women with induction of labor to those in spontaneous labor. This is problematic, because at any point in the management of the woman with a term gestation, the clinician has the choice between induction of labor and expectant management, not spontaneous labor. Expectant management of the pregnancy involves nonintervention at any particular point in time and allowing the pregnancy to progress to a future gestational age. Thus, women undergoing expectant management may go into spontaneous labor or may require indicated induction of labor at a future gestational age., Objectives: The Stanford-UCSF Evidence-Based Practice Center examined the evidence regarding four Key Questions: What evidence describes the maternal risks of elective induction versus expectant management? What evidence describes the fetal/neonatal risks of elective induction versus expectant management? What is the evidence that certain physical conditions/patient characteristics are predictive of a successful induction of labor? How is a failed induction defined?, Methods: We performed a systematic review to answer the Key Questions. We searched MEDLINE(1966-2007) and bibliographies of prior systematic reviews and the included studies for English language studies of maternal and fetal outcomes after elective induction of labor. We evaluated the quality of included studies. When possible, we synthesized study data using random effects models. We also evaluated the potential clinical outcomes and cost-effectiveness of elective induction of labor versus expectant management of pregnancy labor at 41, 40, and 39 weeks' gestation using decision-analytic models., Results: Our searches identified 3,722 potentially relevant articles, of which 76 articles met inclusion criteria. Nine RCTs compared expectant management with elective induction of labor. We found that overall, expectant management of pregnancy was associated with an approximately 22 percent higher odds of cesarean delivery than elective induction of labor (OR 1.22, 95 percent CI 1.07-1.39; absolute risk difference 1.9, 95 percent CI: 0.2-3.7 percent). The majority of these studies were in women at or beyond 41 weeks of gestation (OR 1.21, 95 percent CI 1.01-1.46). In studies of women at or beyond 41 weeks of gestation, the evidence was rated as moderate because of the size and number of studies and consistency of the findings. Among women less than 41 weeks of gestation, there were three trials which reported no difference in risk of cesarean delivery among women who were induced as compared to expectant management (OR 1.73; 95 percent CI: 0.67-4.5, P=0.26), but all of these trials were small, non-U.S., older, and of poor quality. When we stratified the analysis by country, we found that the odds of cesarean delivery were higher in women who were expectantly managed compared to elective induction of labor in studies conducted outside the U.S. (OR 1.22; 95 percent CI 1.05-1.40) but were not statistically different in studies conducted in the U.S. (OR 1.28; 95 percent CI 0.65-2.49). Women who were expectantly managed were also more likely to have meconium-stained amniotic fluid than those who were electively induced (OR 2.04; 95 percent CI: 1.34-3.09). Observational studies reported a consistently lower risk of cesarean delivery among women who underwent spontaneous labor (6 percent) compared with women who had an elective induction of labor (8 percent) with a statistically significant decrease when combined (OR 0.63; 95 percent CI: 0.49-0.79), but again utilized the wrong control group and did not appropriately adjust for gestational age. We found moderate to high quality evidence that increased parity, a more favorable cervical status as assessed by a higher Bishop score, and decreased gestational age were associated with successful labor induction (58 percent of the included studies defined success as achieving a vaginal delivery anytime after the onset of the induction of labor; in these instances, induction was considered a failure when it led to a cesarean delivery). In the decision analytic model, we utilized a baseline assumption of no difference in cesarean delivery between the two arms as there was no statistically significant difference in the U.S. studies or in women prior to 41 0/7 weeks of gestation. In each of the models, women who were electively induced had better overall outcomes among both mothers and neonates as estimated by total quality-adjusted life years (QALYs) as well as by reduction in specific perinatal outcomes such as shoulder dystocia, meconium aspiration syndrome, and preeclampsia. Additionally, induction of labor was cost-effective at $10,789 per QALY with elective induction of labor at 41 weeks of gestation, $9,932 per QALY at 40 weeks of gestation, and $20,222 per QALY at 39 weeks of gestation utilizing a cost-effectiveness threshold of $50,000 per QALY. At 41 weeks of gestation, these results were generally robust to variations in the assumed ranges in univariate and multi-way sensitivity analyses. However, the findings of cost-effectiveness at 40 and 39 weeks of gestation were not robust to the ranges of the assumptions. In addition, the strength of evidence for some model inputs was low, therefore our analyses are exploratory rather than definitive., Conclusions: Randomized controlled trials suggest that elective induction of labor at 41 weeks of gestation and beyond may be associated with a decrease in both the risk of cesarean delivery and of meconium-stained amniotic fluid. The evidence regarding elective induction of labor prior to 41 weeks of gestation is insufficient to draw any conclusion. There is a paucity of information from prospective RCTs examining other maternal or neonatal outcomes in the setting of elective induction of labor. Observational studies found higher rates of cesarean delivery with elective induction of labor, but compared women undergoing induction of labor to women in spontaneous labor and were subject to potential confounding bias, particularly from gestational age. Such studies do not inform the question of how elective induction of labor affects maternal or neonatal outcomes. Elective induction of labor at 41 weeks of gestation and potentially earlier also appears to be a cost-effective intervention, but because of the need for further data to populate these models our analyses are not definitive. Despite the evidence from the prospective, RCTs reported above, there are concerns about the translation of such findings into actual practice, thus, there is a great need for studying the translation of such research into settings where the majority of obstetric care is provided.

Background: Although paediatric high grade gliomas resemble their adult counterparts in many ways, there appear to be distinct clinical and biological differences. One important factor hampering the development of new targeted therapies is the relative lack of cell lines derived from childhood glioma patients, as it is unclear whether the well-established adult lines commonly used are representative of the underlying molecular genetics of childhood tumours. We have carried out a detailed molecular and phenotypic characterisation of a series of paediatric high grade glioma cell lines in comparison to routinely used adult lines., Principal Findings: All lines proliferate as adherent monolayers and express glial markers. Copy number profiling revealed complex genomes including amplification and deletions of genes known to be pivotal in core glioblastoma signalling pathways. Expression profiling identified 93 differentially expressed genes which were able to distinguish between the adult and paediatric high grade cell lines, including a number of kinases and co-ordinated sets of genes associated with DNA integrity and the immune response., Significance: These data demonstrate that glioma cell lines derived from paediatric patients show key molecular differences to those from adults, some of which are well known, whilst others may provide novel targets for evaluation in primary tumours. We thus provide the rationale and demonstrate the practicability of using paediatric glioma cell lines for preclinical and mechanistic studies.

Case Description: A 21-month-old spayed female Border Collie was examined because of progressive right forelimb lameness, signs of pain, and subcutaneous edema. The dog lived in a fenced yard in Tampa, Fla, that contained a small area of marshy terrain., Clinical Findings: The subcutis and intermuscular fascia contained multiple cystic cavities filled with larval cestodes (plerocercoids or spargana) and cloudy red fluid. Parasites were identified morphologically and by DNA sequence analysis as pseudophyllidean cestodes, most likely Sparganum proliferum. The dog developed a progressively worsening fever, dyspnea, mature neutrophilia, and hypoproteinemia. Septic pleuritis and peritonitis complicated the later stages of the disease., Treatment and Outcome: Treatment with praziquantel, fenbendazole, and nitazoxanide failed to control the proliferation and dissemination of larval cestodes. The dog was euthanatized after 133 days of treatment. At necropsy, numerous parasitic tissue cysts were present in the subcutis and intermuscular fascia; these cysts were most abundant in the soft tissues of the forelimbs and cervical musculature. The pleural and peritoneal cavities contained multiple larval cestodes and were characterized by neutrophilic inflammation and secondary bacterial infection., Clinical Relevance: Findings indicated that clinical signs associated with proliferative sparganosis in dogs may be rapidly progressive and that the condition may be refractory to antiparasitic treatment. Veterinarians should be aware of this zoonotic, water-borne agent.

Hepatozoon (H.) americanum and H. canis are the etiological agents of canine hepatozoonosis, a disease that is found worldwide and is also prevalent in the southeastern United States. Current laboratory diagnosis of canine hepatozoonosis caused by H. americanum is usually dependent on visual identification of Hepatozoon "onion skin cysts" in muscle biopsies, an approach that requires invasive sampling and can result in false negatives. We have developed a diagnostic method for detection of Hepatozoon spp. DNA that integrates nucleic acid extraction with extensive agitation to maximize DNA extraction efficiency. The DNA extracted from canine EDTA-whole blood is subjected to real-time PCR, and fluorescence resonance energy transfer (FRET) probes detect a signature polymorphism in the amplified DNA. This PCR method amplifies a fragment of the Hepatozoon 18S rDNA gene, detects as few as 7 genomic copies of Hepatozoon spp. per ml of blood with high specificity, and differentiates between H. americanum and H. canis amplicons. A surprising 300-fold increase of H. americanum 18S rDNA targets occurred during 3-0 days of storage of positive blood specimens. Examination of 614 EDTA-blood samples submitted mostly from the southeastern Unites States from dogs with suspected hepatozoonosis identified H. americanum in 167 samples (27.2%). An additional 14 samples (2.3%) were positive for H. canis, and 14 samples (2.3%) were positive for both H. americanum and H. canis. These results suggest that the Hepatozoon spp. 18S rDNA quantitative PCR may be a valuable tool that can improve diagnosis and therapy of canine hepatozoonosis.

We investigated the experimental susceptibility and natural exposure of raccoons (Procyon lotor) to five tick-borne pathogens of human and veterinary importance, Ehrlichia canis, E. chaffeensis, E. ewingii, Anaplasma phagocytophilum (ApVariant 1 and Ap-ha HGE-1 strains), and Borrelia lonestari. Infections were assessed by polymerase chain reaction (PCR), indirect fluorescent antibody (IFA) testing, and/or culture isolation methods for at least 30 days postinoculation (DPI). Two E. chaffeensis-inoculated raccoons seroconverted and were transiently PCR positive. One raccoon was culture positive. Laboratory raised Amblyomma americanum nymphs fed on a third infected raccoon failed to become infected. Two A. phagocytophilum (HGE-1)-inoculated raccoons became PCR positive and seroconverted. Both remained positive for at least 74 DPI. In contrast, raccoons inoculated with A. phagocytophilum (Ap-Variant 1) were only transiently PCR positive and only seroconverted with low titers. No evidence of infection was observed for E. ewingii- and B. lonestari-inoculated raccoons. Only one E. canis-inoculated raccoon was PCR positive 3 DPI. Serologic testing of wild raccoons from five populations (3 infested with ticks) in Georgia and Florida showed antibodies reactive with E. chaffeensis in the 3 tick-infested populations (range of 30%-46%), E. canis in the same three populations (8%-23%), A. phagocytophilum in a single raccoon from Florida (12%), and Borrelia spp. in all 5 populations (8%-53%). All raccoons were PCR negative for tick-borne pathogens. These data suggest that raccoons are likely not important reservoirs of E. canis, E. ewingii, or B. lonestari. However, raccoons are experimentally susceptible and naturally exposed to E. chaffeensis, and these data support the previous finding that raccoons may be involved in the natural history of A. phagocytophilum.

Hepatozoon americanum is a protozoan that causes American canine hepatozoonosis (ACH) in the southern United States; Hepatozoon canis, the causative agent of canine hepatozoonosis in Africa, Asia, Europe, and South America, has not previously been definitively identified in dogs in the United States. To characterize the diversity of Hepatozoon spp. in domestic dogs from Oklahoma, blood samples collected from dogs residing in an endemic area of the state, clinical cases presented to veterinarians with symptoms of ACH, and dogs housed at a local shelter were evaluated by a nested PCR designed to amplify a variable region of the 18S rRNA gene of blood ampicomplexa, including Hepatozoon spp. Hepatozoon sequences recovered from a dog from an area where ACH is endemic, from clinically ill dogs, and from one shelter dog most closely resembled H. americanum. However, two other shelter dogs had evidence of infection with H. canis or a closely related organism. A subsequent review of real-time PCR results from the Molecular Diagnostics Laboratory at Auburn University revealed that the majority of samples submitted from dogs from across the United States which tested positive for Hepatozoon spp. had H. americanum. However, some submissions were also found which contained DNA sequence of H. canis. Mixed H. americanum and H. canis-like infections also were detected. Our data suggest that H. americanum, H. canis, as well as H. canis-like organisms are present and may cause disease in dogs in the southern U.S.

Hepatozoon americanum cystozoites from experimentally infected, laboratory-raised rodents were fed to a Hepatozoon-free dog. Gamonts were detected by examination of blood smear 42 and 56 days post-exposure. PCR analysis of blood was positive for the 18S rRNA Hepatozoon gene on days gamonts were demonstrated. Meronts were detected histologically in a skeletal muscle biopsy 90 days after ingestion of cystozoites. Sequencing confirmed that the parasite in the dog was H. americanum. Xenodiagnosis was conducted by replete feeding of Ambylomma maculatum larvae on the dog; 40 days after detachment, sporulated oocysts were recovered from recently molted nymphs.

An Ehrlichia sp. (Panola Mountain [PM] Ehrlichia sp.) closely related to Ehrlichia ruminantium was recently detected in a domestic goat experimentally infested with lone star ticks (LSTs, Amblyomma americanum) collected from Georgia, USA. The infected goat exhibited pyrexia and mild clinical pathologic abnormalities consistent with ehrlichiosis. At least two other Ehrlichia species (Ehrlichia chaffeensis and Ehrlichia ewingii) are maintained in nature by a cycle involving LSTs as the primary vector and white-tailed deer (Odocoileus virginanus) as a known or suspected reservoir. To investigate the possibility that white-tailed deer are potential hosts of the PM Ehrlichia sp., whole blood samples collected from 87 wild deer from 2000 to 2002 were screened with a species-specific nested PCR assay targeting the citrate synthase gene. In addition, two laboratory-raised white-tailed deer fawns were each infested with 120 wild-caught LST adults from Missouri, USA, and blood samples were periodically collected and tested for the PM Ehrlichia sp. Of 87 deer tested from 20 locations in the southeastern United States, three (3%) deer from Arkansas, North Carolina, and Virginia were positive for the PM Ehrlichia sp. Wild-caught ticks transmitted the PM Ehrlichia sp. to one of two deer fawns, and colony-reared nymphal LSTs acquired the organism from the deer, maintained it transstadially as they molted to adults, and transmitted the PM Ehrlichia sp. to two naïve fawns. These findings indicate that white-tailed deer are naturally and experimentally susceptible to infection with an Ehrlichia sp. closely related to E. ruminantium and are able to serve as a source of infection to LSTs.

Heart failure patients have abnormal cardiac high energy phosphate metabolism, the explanation for which is unknown. Patients with heart failure also have elevated plasma free fatty acid (FFA) concentrations. Elevated FFA levels are associated with increased cardiac mitochondrial uncoupling proteins (UCPs), which, in turn, are associated with decreased mitochondrial respiratory coupling and low cardiac efficiency. Here, we determined whether increased mitochondrial UCP levels contribute to decreased energetics in the failing heart by measuring UCPs and respiration in mitochondria isolated from the viable myocardium of chronically infarcted rat hearts and measuring efficiency (hydraulic work/O(2) consumption) in the isolated, working rat heart. Ten weeks after infarction, cardiac levels of UCP3 were increased by 53% in infarcted, failing hearts that had ejection fractions less than 45%. Cardiac UCP3 levels correlated positively with non-fasting plasma FFAs (r=0.81; p<0.01). Mitochondria from failing hearts were less coupled than those from control hearts, as demonstrated by the lower ADP/O ratio of 1.9+/-0.1 compared with 2.5+/-0.2 in controls (p<0.05). The decreased ADP/O ratio was reflected in an efficiency of 14+/-2% in the failing hearts when perfused with 1 mM palmitate, compared with 20+/-1% in controls (p<0.05). We conclude that failing hearts have increased UCP3 levels that are associated with high circulating FFA concentrations, mitochondrial uncoupling, and decreased cardiac efficiency. Thus, respiratory uncoupling may underlie the abnormal energetics and low efficiency in the failing heart, although whether this is maladaptive or adaptive would require direct investigation.

Laboratory-raised cotton rats (Sigmodon hispidus), outbred white mice (Mus musculus), and C57BL/6J-Lystbg-J/J mice (M. musculus) that were administered approximately 50 sporulated oocysts of Hepatozoon americanum (AF176836) by gavage developed inflammatory lesions containing parasitic cystozoites in cardiac and skeletal muscle, kidney, and lung. Sprague-Dawley rats (Rattus norvegicus) similarly exposed showed no evidence of infection. Cystozoites were first detected by histopathologic examination four weeks after exposure to oocysts. Globular, PAS-positive material accumulated around the cystozoites as the duration of infection lengthened. Nested PCR analysis of tissues collected 16 weeks post-exposure was positive for the 18S rRNA Hepatozoon sp. gene and the DNA sequence of the fragment amplified was 99.6% and 99.8% identical to H. americanum sequences previously reported from naturally-infected dogs (AF176836 and AY864676, respectively). Merogonous and gamontogonous stages of the parasite were not detected in any of the cystozoite-infected rodents.

In the summer of 2006, an Amblyomma americanum tick was removed from a woman in central North Carolina, who subsequently developed a rash at the site of tick attachment. When examined by polymerase chain reaction (PCR) for Borrelia, Anaplasma, Ehrlichia, Babesia, Rickettsia, and Bartonella DNA, only the Rickettsia primers generated an amplicon, which was identified as "R. amblyommii" by sequencing. To our knowledge, this is the first case in which R. amblyommii was temporally associated with a rash.

Adult male ticks have been shown capable of experimental acquisition and transmission of tick-borne pathogens without requiring a molt. To determine the ecological relevance of this intrastadial transmission route, we evaluated the extent to which actively feeding male Rhipicephalus sanguineus move naturally between co-housed infested dogs. Dogs (n=4) were infested with single color-coded ticks, individually housed in tick-confinement cages for 48 h while infestations established, and then each dog examined and the ticks present counted. Dogs were then co-housed in a large, group pen for an additional 7 (study 1) or 5 (study 2) days. In the first study, sex ratios were adjusted to encourage migration, with two dogs receiving predominantly female ticks and two dogs receiving all male ticks. In the second study, each dog received a ratio of ticks that parallels that found in natural infestations (4:1 male to female). Results showed that ticks readily migrated between infested, co-housed dogs. Rates of immigration, defined as the percentage of ticks previously attached to one dog that moved onto another dog, ranged from 0 to 46% (mean=31.1% study 1; 9.4% study 2). Emigration rates, defined as the number of ticks initially infesting one dog that moved to another dog, averaged 35.2% in study 1 and 10.8% in study 2 (3.6-67.6%). Movement of adult ticks between dogs represents a naturally occurring form of interrupted feeding, a strategy which has been shown to shorten the feeding time necessary to allow transmission of pathogens. In ticks that readily detach from one host and reattach to a second host to resume feeding, replication of any pathogens present has already been initiated and therefore the same delay in transmission seen in ticks attached to a host for the first time may not occur.

Eighteen of 31 (58%) cotton rats (Sigmodon hispidus) and 8 of 24 (33.3%) white-footed mice (Peromyscus leucopus) that were wild-trapped from 4 American canine hepatozoonosis endemic sites in Oklahoma were infected with Hepatozoon species. The predilection organ for merogony of the Hepatozoon species in cotton rats was the liver. Meronts were not detected in any of the white-footed mice. A 488 bp DNA fragment that includes a variable region of the 18S rRNA Hepatozoon gene amplified from blood or tissue of these infected animals. Sequences from eight cotton rats were 100% identical to each other as were sequences from three white-footed mice 100% identical to each other. The cotton rat sequence and the white-footed mouse sequence were 98.8% identical, differing in 6 bp of the 488 bp fragment. The DNA sequence from cotton rats was 97.7% identical to a Hepatozoon sp. described in a large bandicoot rat from Thailand and 97.5% identical to a Hepatozoon sp. in a bank vole from Brazil. The sequence from white-footed mice was 98.6% identical to the bandicoot rat sequence and 98.4% identical to the bank vole sequence. However, the sequences were only 90.6% (cotton rat) and 91.4% (white-footed mouse) identical to H. americanum. These findings suggest that the rodents are obligate intermediate hosts for distinct Hepatozoon spp., but not H. americanum.

Purpose: Epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase overexpressed in a variety of human malignancies, against which targeted therapies have shown efficacy in lung and brain tumors. Clinical responses to EGFR inhibitors have been found to be highly dependent on the presence of activating mutations, whereas gene amplification, downstream activation of Akt, and abnormalities in PTEN are also reported predictive factors. We sought to evaluate these variables in pediatric renal tumors., Experimental Design: We screened a series of 307 pediatric renal tumors for EGFR expression by immunohistochemistry and gene amplification by chromogenic in situ hybridization. In identifying a striking predilection for certain tumor types, we further analyzed the clear cell sarcomas of the kidney (CCSK) for mutations in EGFR and PTEN., Results: Although only 23 of 177 (13.0%) nonanaplastic Wilms' tumors were EGFR positive, 4 of 11 (36.4%) anaplastic tumors showed receptor overexpression. In addition, 5 of 9 (55.6%) mesoblastic nephromas and 12 of 12 (100%) CCSKs were strongly immunoreactive for EGFR. In studying the CCSKs in more detail, we identified gene amplification in 1 of 12 (8.3%) cases and a somatic T790M EGFR mutation in a further case. These two samples additionally harbored mutations in PTEN. Downstream pathway activation, as assayed by phosphorylated Akt expression, was observed in 8 of 12 (66.7%) cases., Conclusions: Together, these data show dysregulation of the EGFR pathway at multiple levels in CCSKs. Identification of factors predictive of poor response to targeted therapy, including the drug resistance T790M mutation, may provide a rationale for upfront trials with irreversible inhibitors of EGFR in children with these tumors.

Purpose: The most well established molecular markers of poor outcome in Wilms' tumor are loss of heterozygosity at chromosomes 1p and/or 16q, although to date no specific genes at these loci have been identified. We have previously shown a link between genomic gain of chromosome 1q and tumor relapse and sought to further elucidate the role of genes on 1q in treatment failure., Experimental Design: Microarray-based comparative genomic hybridization identified a microamplification harboring a single gene (CACNA1E) at 1q25.3 in 6 of 76 (7.9%) Wilms' tumors, correlating with a shorter relapse-free survival (P = 0.0044, log-rank test). Further characterization of this gene was carried out by measuring mRNA and protein expression as well as stable transfection of HEK293 cells., Results: Overexpression of the CACNA1E transcript was associated with DNA copy number (P = 0.0204, ANOVA) and tumor relapse (P = 0.0851, log-rank test). Immunohistochemistry against the protein product Ca(V)2.3 revealed expression localized to the apical membrane in the distal tubules of normal kidney but not to the metanephric blastemal cells of fetal kidney from which Wilms' tumors arise. Nuclear localization in 99 of 160 (61.9%) Wilms' tumor cases correlated with a reduced relapse-free survival, particularly in cases treated with preoperative chemotherapy (P = 0.009, log-rank test). Expression profiling of stably transfected HEK293 cells revealed specific up-regulation of the immediate early response genes EGR1/EGR2/EGR3 and FOS/FOSB, mediated by activation of the MEK/ERK5/Nur77 pathway., Conclusions: These data identify a unique genetic aberration with direct clinical relevance in Wilms' tumor relapse and provide evidence for a potential novel mechanism of treatment resistance in these tumors.

Most Wilms' tumors are of low stage, favorable histology, and have a high likelihood of cure with current multimodal therapy. Despite this, there remains a group of patients whose tumors recur for whom intensive salvage regimens result in survival of only 50%. Fitting a Cox proportional hazards model to microarray-based comparative genomic hybridization (aCGH) data on 68 Wilms' tumor samples, we identified a significant correlation between increased copy number at chromosome 15q26.3 insulin-like growth factor I receptor (IGFIR) and tumor relapse (adjusted P = 0.014). Wilms' tumors (13%) exhibited a low-level gain corresponding to three to four copies of the gene by aCGH analysis, 9 of 10 of which exhibited high IGFIR mRNA levels. Although IGFIR protein expression was restricted to the epithelial cells of fetal kidney and Wilms' tumors in most cases, 12% of tumors were also found to express IGFIR in the blastemal compartment. Blastemal IGFIR protein expression was associated with an increased copy number and a shorter relapse-free survival time (P = 0.027, log-rank test). In addition to the membrane localization, IGFIR was localized to the perinuclear region of the blastemal cells in 6% of Wilms' tumors. These data provide evidence that an increase in IGFIR gene copy number results in aberrant expression in the blastemal compartment of some Wilms' tumors and is associated with an adverse outcome in these patients. These findings suggest the possibility of use of targeted agents in the therapy of these children.

Borrelia lonestari is considered a putative agent of southern tick-associated rash illness (STARI) and is known to occur naturally only in lone star ticks (Amblyomma americanum) and white-tailed deer (Odocoileus virginianus). We used a low passage isolate of B. lonestari (LS-1) to inoculate white-tailed deer, C3H mice, Holstein cattle, and beagles. Animals were monitored via examination of Giemsa and acridine orange stained blood smears, polymerase chain reaction (PCR), indirect fluorescent antibody (IFA) test, and/or culture isolation. Spirochetes were visualized in blood smears of both deer on days post-inoculation (DPI) 6, 8, 12 and one deer on DPI 15. Whole blood collected from deer tested PCR positive starting on DPI 4 and remained positive as long as DPI 28. Both deer developed antibody titers of >64, with a maximum IFA titer of 1024. The organism was reisolated from the blood of both deer on DPI 6 and one deer on DPI 12. All isolation attempts from mice, calves, or dogs were negative, although one of seven mice was transiently PCR positive. Mice and dogs developed an IFA titer > or =64, while calves lacked a detectable antibody response. These preliminary experimental infection trials show that white-tailed deer are susceptible to infection with B. lonestari and develop a spirochetemia following needle-inoculation, while C3H mice, calves, and dogs do not. Results suggest that deer may serve as a vertebrate reservoir host. Tick transmission studies are needed to confirm that this organism can be maintained in a natural cycle involving deer and A. americanum.

We investigated the effect of exposing deer to multiple strains of Ehrlichia chaffeensis that differed in number of tandem repeats in either the variable-length PCR target (VLPT) gene or 120 kDa antigen gene. We hypothesized that infection with one strain would provide immunity to infection with other strains of E. chaffeensis. All deer initially exposed to strain A (604-2) became PCR and culture positive by 10 days post-infection (DPI). Three deer infected with strain A and subsequently inoculated with strain B (623-4) became infected with strain B. Two deer infected with strain A and subsequently inoculated with strain C (125B) became infected with strain C. Of three deer, each infected with strain B and subsequently inoculated with strain C, one was PCR positive for strain C. Of three deer previously inoculated with both strains A and B, and subsequently inoculated with strain C, one showed delayed evidence of strain C. Western blot analysis demonstrated that deer sera reacted differently to antigens from each exposed strain. A complementary in vitro study demonstrated that exposure to two strains differing in VLPT repeats may lead to co-infection of DH82 cells. These results complement a previous study and further show that deer can become sequentially infected with up to three strains of E. chaffeensis. This suggests that competitive exclusion, a phenomenon described in related organisms such as Anaplasma marginale whereby infection with one strain precludes subsequent infection by a second, distinct strain of the same species, may not occur with E. chaffeensis.