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Background Brown dog ticks, Rhipicephalus sanguineus sensu lato (s.l.), have spread globally and are an important vector of multiple pathogens affecting both dogs and humans. The control of these ticks on dogs and human dwellings often relies on synthetic pyrethroids, a chemical class of acaricides that targets the voltage-gated sodium channel (VGSC) protein of arthropod nerve cells, causing influxes of sodium and ultimately, paralysis. Invasive Rhipicephalus sanguineus s.l. ticks in the Americas can be broadly grouped into two main lineages: temperate and tropical ( Rh. sanguineus sensu stricto (s.s.) and Rh. linnaei , respectively). Phenotypic resistance to synthetic pyrethroids in a verified Rh. sanguineus s.s. population has only been reported in the state of Rio Grande do Sul, Brazil. The objectives of our study were to 1) screen a small number ( n  = 10) of Rh. sanguineus s.s. from Brazil to check for VGSC gene mutations known to be associated with resistance to synthetic pyrethroids in ticks and other arthropods, and 2) provide molecular confirmation that all ticks were Rh. sanguineus s.s. Methods We used next-generation DNA sequencing methods to analyze the VGSC gene and mitochondrial loci ( 12S , 16S , COI ) of 10 brown dog ticks sampled from a stray dog in the Restinga sub-district of Porto Alegre, Rio Grande do Sul, Brazil. The progeny of other ticks from this animal had previously been shown to display a low level of resistance to deltamethrin. Results Analysis of mitochondrial genes confirmed these ticks were Rh. sanguineus s.s. We identified two known resistance mutations in domain II segments 4 and 5 of the VGSC gene (C190A and G215T). These specific mutations have not been reported previously in any brown dog tick lineages from the Americas, and this is the first case of VGSC mutations described from ticks confirmed to be Rh. sanguineus s.s. using genetic analyses. Conclusions The discovery of these mutations in Rh. sanguineus s.s. is important for the effective management of ticks on dogs in Brazil and other countries where brown dog tick infestations occur. Graphical Abstract

All human cases of Valley fever are acquired through environmental exposure, so surveillance and characterization of the pathogen in soil are critical for risk mitigation efforts. Current databases are biased toward human clinical isolates, and little is known about the genomics of environmental strains of Coccidioides posadasii . In this study, we designed, tested, and validated a probe enrichment system that amplifies trace DNA in a complex sample. Sequenced DNA can be used to link environmental exposure with human cases, directing public health agencies to interventions that limit human exposure. This use case was demonstrated in this study, as trace DNA trapped on air filters was linked to a fatal case of primate Coccidioidomycosis at a site in Arizona. The probe enrichment system described in this study represents a powerful tool to better understand the genomic composition of environmental C. posadasii strains, which can aid in public health investigations.

Background Amblyomma variegatum threatens the Caribbean cattle industry owing to its role as a vector for Ehrlichia ruminantium , the obligate intracellular bacterium that causes heartwater disease, an economically important and potentially fatal ruminant disease. Amblyomma variegatum is also a public health concern as a vector for Rickettsia africae , the causative agent of African tick-bite fever. Efforts to eradicate A. variegatum on Caribbean islands are ongoing to protect cattle from disease and prevent the spread of the vector and diseases to the American mainland. However, reinfestations often occur, possibly owing to the maintenance of ticks by non-cattle hosts that escape treatment. St. Croix in the US Virgin Islands has experienced such eradication challenges. Methods To determine whether persistence of A. variegatum populations on non-cattle hosts contributes to cattle reinfestation on St. Croix, we analyzed 1 non-attached A. variegatum adult female collected from a human and 14 questing adult females collected via cloth dragging along vegetation transects in Lower Love, St. Croix, during 2023. We conducted host bloodmeal analysis by obtaining vertebrate cytochrome c oxidase subunit I ( COI ) sequences from A. variegatum DNA extracts using COI -targeted DNA capture and enrichment to identify the most recent host bloodmeal. We also screened tick DNA extracts for E. ruminantium and Rickettsia DNA using pCS20 Sol1 and PanR8 qPCR, respectively. Rickettsia species identification was determined using ompA polymerase chain reaction (PCR) and Sanger sequencing, followed by phylogenetic analysis. Results We identified vertebrate COI sequences for the genera Capra and Canis in two A. variegatum ticks. Although E. ruminantium was not detected, DNA from R. africae was present in all 15 ticks. Conclusions These results suggest that goats, and possibly canines, may serve as alternative hosts for A. variegatum on St. Croix, US Virgin Islands, which complicates eradication efforts focused entirely on the treatment of cattle. Fortunately, E. ruminantium was not identified in any ticks. However, R. africae was ubiquitous, which may be of concern for public health. Graphical Abstract

In recent years, an increasing diversity of species has been recognized within the family Francisellaceae. Unfortunately, novel isolates are sometimes misnamed in initial publications or multiple sources propose different nomenclature for genetically highly similar isolates. Thus, unstructured and occasionally incorrect information can lead to confusion in the scientific community. Historically, detection of Francisella tularensis in environmental samples has been challenging due to the considerable and unknown genetic diversity within the family, which can result in false positive results. We have assembled a comprehensive collection of genome sequences representing most known Francisellaceae species/strains and restructured them according to a taxonomy that is based on phylogenetic structure. From this structured dataset, we identified a small number of genomic regions unique to F. tularensis that are putatively suitable for specific detection of this pathogen in environmental samples. We designed and validated specific PCR assays based on these genetic regions that can be used for the detection of F. tularensis in environmental samples, such as water and air filters.

Plague is a rapidly progressing and often fatal zoonotic disease caused by the bacterium Yersinia pestis. This pathogen has impacted humans for millennia, especially during three major pandemics that resulted in over 200 million deaths. During the third pandemic, plague was involved in a global spreading event, and it is now ecologically established on five continents, including North America. Although plague has historically had major implications for humans, in North America it is now primarily a disease of rodents and their associated fleas. Plague cycles between amplifying events where disease transmission is rapid (epizootics), and low-level persistence cycles (enzootics). It is during epizootic events that large die offs of infected rodent species are documented, and infected fleas can feed on humans. This results in the most common clinical presentation of the disease in humans, bubonic plague. Bubonic plague is acquired via flea vectors and, if left untreated, has a 40-60% mortality rate. Bubonic plague can progress to the less common but more fatal form of the disease, pneumonic plague, which approaches 100% mortality within 1-5 days of disease onset (8). Although outbreaks of sylvatic plague in the southwestern US are well documented, the mechanisms by which Y. pestis persists are unknown. Sylvatic plague is defined as a flea-borne disease in wild rodents. Sampling efforts to detect this disease during inter-epizootic periods yield low or no detection of Y. pestis. Previous studies examining multi-locus variable number tandem repeat loci (MLVA) in Y. pestis have shown local differentiation when examined at one time point (7), but data needed to sufficiently analyze persistence through time for this pathogen is not readily available. This study examines Y. pestis genomic diversity across time (2001-2020) and space (nine locations) near Flagstaff, Arizona, USA and utilizes archived flea DNA extracts that were collected for public health purposes. Therefore, due to a limited number of fleas collected at each site, not all sites had additional fleas that could be used for culturing of Y. pestis. Complex DNA extracts obtained from whole fleas are poor candidates for whole genome sequencing of Y. pestis due to low burden of pathogen and therefore would produce low coverage of the Y. pestis genome. Therefore, to characterize Y. pestis genomes present in these flea samples, culture methods to obtain pure isolates and targeted Y. pestis DNA capture and enrichment were used and whole genome sequenced. Target enrichment uses of RNA probes to capture the genome of Y. pestis present in the flea DNA extractions. Genomic data from both enrichment and cultured samples revealed two main phylogenetic lineages of Y. pestis located in east and west Flagstaff, each with multiple subclades. In most cases, samples within a (the same?) subclade were found within ~8 kilometers of each other and displayed a limited distribution within the Flagstaff area. However, in one case, closely related Y. pestis genotypes had spread between Williams, and Flagstaff, AZ, revealing the ability to disperse ~77 kilometers within three years. We also found examples of genomes sampled 10 years apart with no identified variation (n=0 SNPs), suggesting Y. pestis can persist with low replication levels through extended periods of time. This study has allowed us to identify local phylogenetic lineages of Y. pestis circulating near Flagstaff, AZ and follow their persistence for two decades.

This study evaluated receipt of HIV prevention services in correctional substance abuse treatment programs and examined their impact on short-term risk behaviors. The authors performed a secondary analysis of the National Treatment Improvement Evaluation Study (NTIES), a prospective cohort study conducted during 1993 to 1995. The sample included 1,223 adult non-HIV-positive inmates, enrolled in nine correctional substance abuse treatment programs. A composite index modeled after the validated Risk Assessment Battery measured HIV risk behavior at treatment entry and at 12-month follow-up. Overall, most inmates received HIV prevention services while in treatment. Controlling for potential confounders, HIV prevention services were significantly associated with reduced risk behavior among inmates who were out of custody at follow-up, but no such association was observed among those still in custody. This analysis suggests that HIV prevention services are beneficial in reducing risk behavior among incarcerated individuals whose discharge is expected in the near future.

Leptospirosis (caused by pathogenic bacteria in the genus Leptospira ) is prevalent worldwide but more common in tropical and subtropical regions. Transmission can occur following direct exposure to infected urine from reservoir hosts, or a urine-contaminated environment, which then can serve as an infection source for additional rats and other mammals, including humans. The brown rat, Rattus norvegicus , is an important reservoir of Leptospira spp. in urban settings. We investigated the presence of Leptospira spp. among brown rats in Boston, Massachusetts and hypothesized that rat population dynamics in this urban setting influence the transportation, persistence, and diversity of Leptospira spp. We analyzed DNA from 328 rat kidney samples collected from 17 sites in Boston over a seven-year period (2016–2022); 59 rats representing 12 of 17 sites were positive for Leptospira spp. We used 21 neutral microsatellite loci to genotype 311 rats and utilized the resulting data to investigate genetic connectivity among sampling sites. We generated whole genome sequences for 28 Leptospira spp. isolates obtained from frozen and fresh tissue from some of the 59 positive rat kidneys. When isolates were not obtained, we attempted genomic DNA capture and enrichment, which yielded 14 additional Leptospira spp. genomes from rats. We also generated an enriched Leptospira spp. genome from a 2018 human case in Boston. We found evidence of high genetic structure among rat populations that is likely influenced by major roads and/or other dispersal barriers, resulting in distinct rat population groups within the city; at certain sites these groups persisted for multiple years. We identified multiple distinct phylogenetic clades of L. interrogans among rats that were tightly linked to distinct rat populations. This pattern suggests L. interrogans persists in local rat populations and its transportation is influenced by rat population dynamics. Finally, our genomic analyses of the Leptospira spp. detected in the 2018 human leptospirosis case in Boston suggests a link to rats as the source. These findings will be useful for guiding rat control and human leptospirosis mitigation efforts in this and other similar urban settings.

Leptospirosis (caused by pathogenic bacteria in the genus Leptospira ) is prevalent worldwide but more common in tropical and subtropical regions. Transmission can occur following direct exposure to infected urine from reservoir hosts, or a urine-contaminated environment, which then can serve as an infection source for additional rats and other mammals, including humans. The brown rat, Rattus norvegicus , is an important reservoir of Leptospira spp. in urban settings. We investigated the presence of Leptospira spp. among brown rats in Boston, Massachusetts and hypothesized that rat population dynamics in this urban setting influence the transportation, persistence, and diversity of Leptospira spp. We analyzed DNA from 328 rat kidney samples collected from 17 sites in Boston over a seven-year period (2016–2022); 59 rats representing 12 of 17 sites were positive for Leptospira spp. We used 21 neutral microsatellite loci to genotype 311 rats and utilized the resulting data to investigate genetic connectivity among sampling sites. We generated whole genome sequences for 28 Leptospira spp. isolates obtained from frozen and fresh tissue from some of the 59 positive rat kidneys. When isolates were not obtained, we attempted genomic DNA capture and enrichment, which yielded 14 additional Leptospira spp. genomes from rats. We also generated an enriched Leptospira spp. genome from a 2018 human case in Boston. We found evidence of high genetic structure among rat populations that is likely influenced by major roads and/or other dispersal barriers, resulting in distinct rat population groups within the city; at certain sites these groups persisted for multiple years. We identified multiple distinct phylogenetic clades of L. interrogans among rats that were tightly linked to distinct rat populations. This pattern suggests L. interrogans persists in local rat populations and its transportation is influenced by rat population dynamics. Finally, our genomic analyses of the Leptospira spp. detected in the 2018 human leptospirosis case in Boston suggests a link to rats as the source. These findings will be useful for guiding rat control and human leptospirosis mitigation efforts in this and other similar urban settings.

The advent of genetic technology has been utilized by creative defense counsel to argue the presence of a preexisting genetic condition as a superseding or contributing cause to a plaintiff's personal injuries. On Aug 12, 2004, the California Court of Appeals in Cruz v. Superior Court of Orange County' affirmed an order authorizing the infant's mother, serving as the guardian ad litem, to submit to genetic testing under California's version of Federal Rules of Civil Procedure (FRCP) Rule 35. Given this social and political environment, defense counsel's attempt to procure a genetic exam under FRCP Rule 35 is likely to be confronted with strong resistance by plaintiff's counsel. Despite the moral challenges it raises and the difficulty in its application, genetics will continue to be pursued, and the law must brace itself to be able to grapple with the multifaceted problems genetics presents.