Ixodes scapularis ticks harbor a variety of microorganisms, including eukaryotes, bacteria and viruses. Some of these can be transmitted to and cause disease in humans and other vertebrates. Others are not pathogenic, but may impact the ability of the tick to harbor and transmit pathogens. A growing number of studies have examined the influence of bacteria on tick vector competence but the influence of the tick virome remains less clear, despite a surge in the discovery of tick-associated viruses. In this study, we performed shotgun RNA sequencing on 112 individual adult I. scapularis collected in Wisconsin, USA. We characterized the abundance, prevalence and co-infection rates of viruses, bacteria and eukaryotic microorganisms. We identified pairs of tick-infecting microorganisms whose observed co-infection rates were higher or lower than would be expected, or whose RNA levels were positively correlated in co-infected ticks. Many of these co-occurrence and correlation relationships involved two bunyaviruses, South Bay virus and blacklegged tick phlebovirus-1. These viruses were also the most prevalent microorganisms in the ticks we sampled, and had the highest average RNA levels. Evidence of associations between microbes included a positive correlation between RNA levels of South Bay virus and Borrelia burgdorferi, the Lyme disease agent. These findings contribute to the rationale for experimental studies on the impact of viruses on tick biology and vector competence.

The mammalian orthoreovirus double-stranded (ds) RNA-binding protein σ3 is a multifunctional protein that promotes viral protein synthesis and facilitates viral entry and assembly. The dsRNA-binding capacity of σ3 correlates with its capacity to prevent dsRNA-mediated activation of protein kinase R (PKR). However, the effect of σ3 binding to dsRNA during viral infection is largely unknown. To identify functions of σ3 dsRNA-binding activity during reovirus infection, we engineered a panel of thirteen σ3 mutants and screened them for the capacity to bind dsRNA. Six mutants were defective in dsRNA binding, and mutations in these constructs cluster in a putative dsRNA-binding region on the surface of σ3. Two recombinant viruses expressing these σ3 dsRNA-binding mutants, K287T and R296T, display strikingly different phenotypes. In a cell-type dependent manner, K287T, but not R296T, replicates less efficiently than wild-type (WT) virus. In cells in which K287T virus demonstrates a replication deficit, PKR activation occurs and abundant stress granules (SGs) are formed at late times post-infection. In contrast, the R296T virus retains the capacity to suppress activation of PKR and does not mediate formation of SGs at late times post-infection. These findings indicate that σ3 inhibits PKR independently of its capacity to bind dsRNA. In infected mice, K287T produces lower viral titers in the spleen, liver, lungs, and heart relative to WT or R296T. Moreover, mice inoculated with WT or R296T viruses develop myocarditis, whereas those inoculated with K287T do not. Overall, our results indicate that σ3 functions to suppress PKR activation and subsequent SG formation during viral infection and that these functions correlate with virulence in mice.

Serpentoviruses are an emerging group of nidoviruses known to cause respiratory disease in snakes and have been associated with disease in other non-avian reptile species (lizards and turtles). This study describes multiple episodes of respiratory disease-associated mortalities in a collection of juvenile veiled chameleons (Chamaeleo calyptratus). Histopathologic lesions included rhinitis and interstitial pneumonia with epithelial proliferation and abundant mucus. Metagenomic sequencing detected coinfection with two novel serpentoviruses and a novel orthoreovirus. Veiled chameleon serpentoviruses are most closely related to serpentoviruses identified in snakes, lizards, and turtles (approximately 40–50% nucleotide and amino acid identity of ORF1b). Veiled chameleon orthoreovirus is most closely related to reptilian orthoreoviruses identified in snakes (approximately 80–90% nucleotide and amino acid identity of the RNA-dependent RNA polymerase). A high prevalence of serpentovirus infection (>80%) was found in clinically healthy subadult and adult veiled chameleons, suggesting the potential for chronic subclinical carriers. Juvenile veiled chameleons typically exhibited a more rapid progression compared to subadults and adults, indicating a possible age association with morbidity and mortality. This is the first description of a serpentovirus infection in any chameleon species. A causal relationship between serpentovirus infection and respiratory disease in chameleons is suspected. The significance of orthoreovirus coinfection remains unknown.

Galbut virus (family Partitiviridae) infects Drosophila melanogaster and can be transmitted vertically from infected mothers or infected fathers with near perfect efficiency. This form of super-Mendelian inheritance should drive infection to 100% prevalence, and indeed, galbut virus is ubiquitous in wild D. melanogaster populations. However, on average, only about 60% of individual flies are infected. One possible explanation for this is that a subset of flies are resistant to infection. Although galbut virus-infected flies appear healthy, infection may be sufficiently costly to drive selection for resistant hosts, thereby decreasing overall prevalence. To test this hypothesis, we quantified a variety of fitness-related traits in galbut virus-infected flies from two lines from the Drosophila Genetic Reference Panel (DGRP). Galbut virus-infected flies had no difference in average lifespan and total offspring production compared to their uninfected counterparts. Galbut virus-infected DGRP-517 flies pupated and eclosed faster than their uninfected counterparts. Some galbut virus-infected flies exhibited altered sensitivity to viral, bacterial, and fungal pathogens. The microbiome composition of flies was not measurably perturbed by galbut virus infection. Differences in phenotype attributable to galbut virus infection varied as a function of fly sex and DGRP strain, and differences attributable to infection status were dwarfed by larger differences attributable to strain and sex. Thus, galbut virus infection does produce measurable phenotypic changes, with changes being minor, offsetting, and possibly net-negative.

Ticks are important vectors of human and animal disease, and outdoor workers are at elevated risk of tickborne diseases (TBDs). This risk is particularly relevant for Indigenous herd workers managing bison (colloquial: buffalo), which are culturally, spiritually, and economically significant. To address this gap, the InterTribal Buffalo Council and Central States Center for Agricultural Safety and Health partnered with Tick Tag Go (University of Nebraska–Lincoln) to provide education on tick identification, prevention, and TBDs at the 2024 Bison Worker Safety & Herd Health Roundtable. Thirty-four participants, including herd managers, workers, and tribal leaders, engaged in the session, with 27 completing a post-presentation survey. The presentation was well received: 96.3% reported gaining new knowledge, 70.4% indicated they were more likely to implement preventive practices, and 74.1% found the material relevant to their needs. Nearly half (44.4%) expressed interest in additional community-based programming. To our knowledge, this is the first report assessing perceptions of ticks and TBDs among Indigenous agricultural workers. Findings highlight both knowledge gaps and receptiveness to education, underscoring the importance of culturally tailored outreach to reduce TBD risk in Indigenous communities.

Purpose: Alpha-gal syndrome (AGS) is an emerging health issue. This syndrome, caused by the bites of ticks, induces allergic reactions to the sugar molecule galactose-α-1,3-galactose after exposure to non-primate mammalian meat and other byproducts. Agricultural workers spend significant time outdoors placing them at an increased risk for tick bites and tick-borne diseases, like AGS. This study aimed to characterize farmers and ranchers’ prior knowledge, symptomology, and diagnostic experiences with AGS. Methods: We conducted a cross-sectional survey of more than 200 farmers and ranchers with a self-reported AGS diagnosis. The survey captured farmers and ranchers’ experiences related to prior knowledge and experience with tick bites and AGS, reported symptoms, and obtaining a diagnosis. Findings: A total of 201 respondents across 26 states participated in the survey, with the majority from Missouri and Oklahoma. We identified four distinct symptom clusters, with the most reported symptoms being abdominal cramping, diarrhea, itchy skin, and nausea. Women more often reported gastrointestinal discomfort, and men were more likely to be in the mild symptom category. On average, participants reported 2.98 medical provider visits before receiving a diagnosis, most being diagnosed by general practitioners and allergists. Conclusions: No previous studies have focused on the symptom and diagnostic experiences of farmers and ranchers with AGS. Capturing such data is essential as these workers may experience unique occupational challenges following AGS diagnosis. The diagnostic experience data support a continuing need to educate and empower AGS patients and providers, especially agricultural workers and providers serving rural communities.