Borrelia miyamotoi in Human-Biting Ticks, United States, 2013-2019.

During 2013-2019, Borrelia miyamotoi infection was detected in 19 US states. Infection rate was 0.5%-3.2%; of B. miyamotoi-positive ticks, 59.09% had concurrent infections. B. miyamotoi is homogeneous with 1 genotype from Ixodes scapularis ticks in northeastern and midwestern states and 1 from I. pacificus in western states.

Ehrlichia muris in Ixodes cookei Ticks, Northeastern United States, 2016-2017.

Ehrlichia muris is an agent of human ehrlichiosis. To determine its geographic spread in the United States, during 2016-2017, we tested 8,760 ticks from 45 states. A distinct clade of E. muris found in 3 Ixodes cookei ticks from the northeastern United States suggests transmission by these ticks in this region.

Differential Resistance of Borrelia burgdorferi Clones to Human Serum-Mediated Killing Does Not Correspond to Their Predicted Invasiveness.

Reservoir host associations have been observed among and within Borrelia genospecies, and host complement-mediated killing is a major determinant in these interactions. In North America, only a subset of Borrelia burgdorferi lineages cause the majority of disseminated infections in humans. We hypothesize that differential resistance to human complement-mediated killing may be a major phenotypic determinant of whether a lineage can establish systemic infection. As a corollary, we hypothesize that borreliacidal action may differ among human subjects. To test these hypotheses, we isolated primary B. burgdorferi clones from field-collected ticks and determined whether the killing effects of human serum differed among those clones in vitro and/or whether these effects were consistent among human sera. Clones associated with human invasiveness did not show higher survival in human serum compared to noninvasive clones. These results indicate that differential complement-mediated killing of B. burgdorferi lineages is not a determinant of invasiveness in humans. Only one significant difference in the survivorship of individual clones incubated in different human sera was detected, suggesting that complement-mediated killing of B. burgdorferi is usually similar among humans. Mechanisms other than differential human complement-mediated killing of B. burgdorferi lineages likely explain why only certain lineages cause the majority of disseminated human infections.

Lactate Dehydrogenase Inhibitors Suppress Borrelia burgdorferi Growth In Vitro.

Borrelia burgdorferi, the causative agent of Lyme disease, has a highly reduced genome and relies heavily on glycolysis for carbon metabolism. As such, established inhibitors of lactate dehydrogenase (LDH) were evaluated in cultures to determine the extent of their impacts on B. burgdorferi growth. Both racemic and enantiopure (AT-101) gossypol, as well as oxamate, galloflavin, and stiripentol, caused the dose-dependent suppression of B. burgdorferi growth in vitro. Racemic gossypol and AT-101 were shown to fully inhibit spirochetal growth at concentrations of 70.5 and 187.5 µM, respectively. Differences between racemic gossypol and AT-101 efficacy may indicate that the dextrorotatory enantiomer of gossypol is a more effective inhibitor of B. burgdorferi growth than the levorotatory enantiomer. As a whole, LDH inhibition appears to be a promising mechanism for suppressing Borrelia growth, particularly with bulky LDH inhibitors like gossypol.

Tick Densities and Infection Prevalence on Coastal Islands in Massachusetts, USA: Establishing a Baseline.

Tick-borne diseases and a tick-induced red meat allergy have become increasingly common in the northeastern USA and elsewhere. At the scale of local communities, few studies have documented tick densities or infection levels to characterize current conditions and provide a baseline for further monitoring. Using the town of Nantucket, MA, as a case study, we recorded tick densities by drag sampling along hiking trails in nature preserves on two islands. Nymphal blacklegged ticks (Ixodes scapularis Say) were most abundant at shadier sites and least common in grasslands and scrub oak thickets (Quercus ilicifolia). Lone star ticks (Amblyomma americanum L.) were common on Tuckernuck Island and rare on Nantucket Island, while both tick species were more numerous in 2021 compared to 2020 and 2022. We tested for pathogens in blacklegged nymphs at five sites over two years. In 2020 and 2021, infection levels among the four Nantucket Island sites averaged 10% vs. 19% for Borrelia burgdorferi, 11% vs. 15% for Babesia microti, and 17% (both years) for Anaplasma phagocytophilum, while corresponding levels were significantly greater on Tuckernuck in 2021. Our site-specific, quantitative approach represents a practical example of how potential exposure to tick-borne diseases can be monitored on a local scale.

White-Tailed Deer Serum Kills the Lyme Disease Spirochete, Borrelia burgdorferi.

Borrelia burgdorferi is a human pathogen vectored by Ixodes ticks and maintained in nature by a suite of competent vertebrate reservoirs. White-tailed deer (WTD) are considered to be noncompetent reservoirs for B. burgdorferi. Sera from other deer species have been found to be borreliacidal, and similar mechanisms could explain the lack of reservoir competence of WTD. Therefore, we determined whether WTD serum can kill B. burgdorferi. Using an in vitro serum sensitivity assay and subculturing of spirochetes, we demonstrated that WTD serum effectively kills B. burgdorferi. The borreliacidal activity of WTD serum likely contributes to the inability of WTD to efficiently harbor and transmit B. burgdorferi.

Kruppel-family zinc finger proteins as emerging epigenetic biomarkers in head and neck squamous cell carcinoma.

BACKGROUND: Krüppel-type zinc finger protein genes located on chromosome 19q13 are aberrantly hypermethylated with high frequency in all anatomic sub-sites of head and neck cancers as well as other epithelial tumours resulting in decreased expression. METHODS: We examined prognostic significance of ZNF154 and ZNF132 expression and DNA methylation in independent patient cohort of about 500 head and neck cancer patients in the Cancer Genome Atlas (TCGA). We also overexpressed these genes in HEK-293 cells, as well as the oral cancer cell line UM-SCC-1. RESULTS: In 20 patients from the TCGA cohort of HNSCC patients where ZNF154 and ZNF132 DNA methylation and RNA expression could be compared in tumor and adjacent normal tissue, there was increased DNA methylation and decreased expression of both ZNF154 and ZNF132 in primary tumours. Low ZNF154 and low ZNF132 expression were associated with shorter overall survival in both head and neck squamous cell carcinoma (HNSCC) and lung adenocarcinoma (LUAC patients). While expression of these proteins in HEK-293 cells produced full-length protein, only truncated copies could be expressed in head and neck cancer cells (UM-SCC-1). The truncated version of ZNF154 protein increased doubling time and reduced cell migration in UM-SCC-1 cancer cells. CONCLUSIONS: Both ZNF132 and ZNF154 represent novel clinically significant biomarkers in head and neck cancer with potential tumour suppressive properties. Future studies will address the underlying molecular mechanisms by which ZNF154 expression in HNSCC contributes to the control of cell growth and migration.

Ixodes scapularis Is the Most Susceptible of the Three Canonical Human-Biting Tick Species of North America to Repellent and Acaricidal Effects of the Natural Sesquiterpene, (+)-Nootkatone.

Ticks are vectors of many human and animal zoonotic disease-causing agents causing significant global health and economic strain. Repellents and acaricides are integral to the human capacity for personal protection from tick bites. Nootkatone, a naturally occurring sesquiterpene found in the Alaskan cedar tree, grapefruit, and other sources, has been documented to be a potent acaricide. Research has also noted repellent effects against some tick species. In this study, our aim was to investigate the effect of synthetic, high-purity (+)-nootkatone on adult Ixodes scapularis, Dermacentor variabilis, and Amblyomma americanum ticks in an in vitro, vertical filter paper bioassay. (+)-nootkatone showed compelling tick repellency, but median effective concentrations (EC50) significantly differed among species. Ixodes scapularis were repelled at very low concentrations (EC50 = 0.87 ± 0.05 µg/cm2). Higher concentrations were required to repel D. variabilis (EC50 = 252 ± 12 µg/cm2) and A. americanum (EC50 = 2313 ± 179 µg/cm2). Significant post-exposure mortality, assessed 24 h after repellency trials, was also observed in I. scapularis but was absent entirely in D. variabilis and A. americanum. These tests demonstrate that nootkatone has a promising dual-action personal protection capacity against adult I. scapularis ticks, warranting further investigation in more natural environments and in the presence of host cues.

A Computer Vision-Based Approach for Tick Identification Using Deep Learning Models.

A wide range of pathogens, such as bacteria, viruses, and parasites can be transmitted by ticks and can cause diseases, such as Lyme disease, anaplasmosis, or Rocky Mountain spotted fever. Landscape and climate changes are driving the geographic range expansion of important tick species. The morphological identification of ticks is critical for the assessment of disease risk; however, this process is time-consuming, costly, and requires qualified taxonomic specialists. To address this issue, we constructed a tick identification tool that can differentiate the most encountered human-biting ticks, Amblyomma americanum, Dermacentor variabilis, and Ixodes scapularis, by implementing artificial intelligence methods with deep learning algorithms. Many convolutional neural network (CNN) models (such as VGG, ResNet, or Inception) have been used for image recognition purposes but it is still a very limited application in the use of tick identification. Here, we describe the modified CNN-based models which were trained using a large-scale molecularly verified dataset to identify tick species. The best CNN model achieved a 99.5% accuracy on the test set. These results demonstrate that a computer vision system is a potential alternative tool to help in prescreening ticks for identification, an earlier diagnosis of disease risk, and, as such, could be a valuable resource for health professionals.

A Borrelia burgdorferi outer surface protein C (OspC) genotyping method using Luminex technology.

Borrelia burgdorferi is an important tickborne human pathogen comprising several strains based on nucleotide sequence of the outer surface protein C (ospC) gene. Detection and characterization of different ospC genotypes is vital for research on B. burgdorferi and the risk it poses to humans. Here we present a novel, multiplex assay based on Luminex xMAP technology for the detection of B. burgdorferi ospC genotypes. The assay has five major steps: amplification of the ospC gene, hydrolyzation of surplus primers and nucleotides, incorporation of biotinylated nucleotides into the template DNA, hybridization to Luminex microspheres, and detection of fluorescent signals corresponding to each ospC genotype. We validated the protocol by comparing results obtained from our method against results from an established ospC genotyping method. This protocol can be used for the characterization of ospC genotypes in B. burgdorferi infected ticks, reservoir hosts, and/or clinical samples.

Human-Biting Ixodes Ticks and Pathogen Prevalence from California, Oregon, and Washington.

From July 2006 through August 2017, a passive surveillance study of Ixodes ticks submitted from California, Oregon, and Washington was conducted by the TickReport program at the University of Massachusetts, Amherst. In total, 549 human-biting Ixodes ticks were submitted comprising both endemic and nonendemic species. We found that 430 endemic ticks were from 3 Ixodes species: Ixodes pacificus, Ixodes spinipalpis, and Ixodes angustus, whereas Ixodes scapularis (n = 111) was the most common species among the 119 nonendemic ticks. The submission peak for nymphal I. pacificus and I. spinipalpis was June, while submission peak for adult I. pacificus and nymphal I. angustus was April and September, respectively. Endemic ticks commonly attached to the lower extremities of their victims, and individuals younger than 9 years old were frequently bitten. The infection prevalence of Borrelia burgdorferi sensu lato, Borrelia miyamotoi, and Anaplasma phagocytophilum in I. pacificus ticks was 1.31%, 1.05%, and 0.52%, respectively, and the prevalence of B. burgdorferi s. l. and A. phagocytophilum in I. spinipalpis ticks was 14.29% and 10.71%, respectively. Furthermore, two species within the B. burgdorferi s. l. complex were detected in West Coast ticks: B. burgdorferi sensu stricto and Borrelia lanei. I. spinipalpis had the highest Borrelia prevalence among endemic ticks, and it was caused exclusively by B. lanei. Borrelia mayonii, Babesia microti, and Ehrlichia muris-like agent were not detected in these endemic ticks. In this study, we show that many nonendemic Ixodes ticks (119/549) are most likely acquired from travel to a different geographic region. We report cases of conventionally recognized nonhuman feeders (I. spinipalpis and I. angustus) parasitizing humans. The highest pathogen prevalence in I. spinipalpis may indicate a larger public health threat than previously thought, and the enzootic life cycle and pathogenicity of B. lanei warrant further study.

Capillary electrophoresis artifact due to eosin: implications for the interpretation of molecular diagnostic assays.

Capillary electrophoresis (CE) is a commonly used tool in the analysis of fluorescently labeled PCR amplification products. We have identified a CE artifact caused by the tissue stain eosin that can complicate the interpretation of CE data. The artifact was detected during routine analysis of a DNA sample isolated from a formalin-fixed, paraffin-embedded tissue sample considered histologically suspicious for a B-cell neoplasm. A standard clinical PCR and CE assay for immunoglobulin heavy chain (IGH) gene rearrangement revealed a weak polyclonal population of rearranged IGH genes and a 71 base peak suspicious for IGH clonality. The spectral properties of the 71 base peak were unusual in that although the dominant fluorescence of the peak was blue, it also fluoresced in green and yellow (blue>green>yellow), raising the suspicion that the peak might represent an artifact. CE analysis of the genomic DNA sample without PCR amplification demonstrated the presence of the 71 base peak, suggesting that the artifact was caused by a contaminant within the DNA sample itself. We demonstrate that eosin, which was used to stain the formalin-fixed tissue during processing, yields a discrete 71 base peak of similar morphology to the contaminant peak on CE analysis. The data suggest that eosin in the fixed tissue was not completely eliminated during nucleic acid extraction, resulting in the artifact peak. We discuss the implications of this potentially common contaminant on the interpretation of CE data and demonstrate that artifacts caused by eosin can be avoided by using more stringent DNA purification steps. Histological dyes may fluoresce, and artifacts from them should be considered when primary peaks contain additional underlying peaks of other colors.

The potential impact of new national guidance on primary prevention of cardiovascular disease in people living with HIV.

INTRODUCTION: Cardiovascular disease (CVD) is the leading cause of death in England and Wales. As people living with HIV (PLWH) age, proactive management of CVD risk factors is crucial. The long-awaited draft guidelines for CVD from the National Institute of Clinical Excellence (NICE) propose lipid modification (with statins) and lifestyle modification for 40-74 year olds with >10% (previously >20%) 10-year risk of CVD using QRISK2. We currently use Framingham so compared 3 CVD risk calculators in our cohort and analyzed the impact of a change in CVD threshold on the proportion of our patients who would need intervention. MATERIALS AND METHODS: Framingham, QRISK2 and JBS3 cardiovascular risk calculators were compared in a group of randomly selected patients. Then, to analyze the impact of a change in primary prevention threshold on our cohort, we interrogated a prospectively collected database to identify all individuals who had a documented Framingham risk assessment and applied the current/proposed thresholds accordingly. We performed the same analysis for the three calculator subgroup (recalculating Framingham risk). Finally we surveyed HIV services in England & Wales regarding their choice of calculator. RESULTS: We compared the 3 CVD risk calculators in 100 patients, see Table 1. CONCLUSIONS: Reducing the threshold for cardiovascular preventative measures to 10% vastly increases the number of patients requiring primary intervention, from two- to fourfold depending on risk calculator used. This may have significant implications, including cost, drug-drug interactions and patient experience, that HIV physicians and general practitioners will need to address, ideally in a coordinated and patient-focused manner.