The etiologic agent of Pacific Coast tick fever (PCTF), a moderately severe tickborne illness that resembles Rocky Mountain spotted fever (RMSF), was first isolated in 1966 from specimens of Dermacentor occidentalis (the Pacific Coast tick) obtained in California. For several decades, this bacterium was identified ambiguously as the unclassified spotted fever group Rickettsia species 364-D, Rickettsia 364, or Rickettsia philipii. However, none of these epithets satisfied criteria of formal bacterial nomenclature. Data developed from mouse serotyping studies performed 45 years ago, and multi-locus sequence typing several decades later, indicated that this bacterium was similar to, but distinct from isolates of Rickettsia rickettsii, the etiological agent of RMSF. We applied an integrative taxonomic approach, combining phenotypic, ecological, and clinical data with whole genome sequencing of 11 contemporary isolates of this pathogen to identify it as a distinct subspecies of R. rickettsii, and propose the name Rickettsia rickettsii subsp. californica subsp. nov.

Spotted fever rickettsiosis is rarely observed in solid organ transplant recipients, and all previously reported cases have been associated with tick bite months to years after transplantation. We describe a kidney transplant recipient in North Carolina, USA, who had a moderately severe Rickettsia parkeri infection develop during the immediate posttransplant period.

Some of the most prevalent arthropod-borne pathogens impacting humans in the United States are transmitted by Ixodes ticks. However, little is known regarding the Rickettsia species that inhabit Ixodes scapularis in the United States. The aim of this study was to screen adult I. scapularis collected in central Oklahoma over an 8-yr period for the presence of tick-borne rickettsial pathogens or potential pathogens. During 2014-2021, 112 adult specimens of I. scapularis were collected from central Oklahoma. Amplicons for Rickettsia spp. were amplified from 53 (47.3%) of the samples. Of the positive ticks, 42 (79.2%) amplicon-positive Rickettsia samples were 100% identical to Rickettsia buchneri, 10 (18.9%) were 100% identical to R. tillamookensis strain Tillamook 23, and 1 (1.9%) specimen showed high identity for Rickettsia amblyommatis. This study highlights the importance of considering Rickettsia-specific assays when assessing Ixodes species ticks for potential pathogens.

We detected the DNA of an Anaplasma bovis-like bacterium in blood specimens from 4 patients from the United States with suspected tickborne illnesses. Initial molecular characterization of this novel agent reveals identity to A. bovis-like bacteria detected in Dermacentor variabilis ticks collected from multiple US states.

In the published version of this article there was an error with the sequence data. Please see the corrected text below: | | | In the abstract | | Upon reanalysis with corrected 16S rRNA sequence, the closest full-length match to non-genome strains in Genbank at the time of this corrigendum was to ‘Candidatus Rickettsia senegalensis’ strain R184 (Genbank accession number OM311169) at 99.7 % identity. | | | In table 1 | | The 16S rRNA column analysis was performed with the chimeric 16S rRNA sequence and is thus in error. The top non-genome strain match and identity is as described for changes to the Abstract. The best match to a complete genome was to R. felis URRWXCal2 (Genbank CP000053) at 99.8 % identity. | | | In phylogeny | | 16S rRNA comparisons are in error and are corrected as for Abstract and Table 1. | | In summary, we have, through antibiotic clearance of Mycoplasma contamination of R. tillamookensis Tillamook 23T, ascertained that corrections need to be made to the published genome record for 16S and 23S rRNA sequences. Genbank records have been updated accordingly. These changes do not affect the status of R. tillamookensis as a distinct species or its phylogenetic placement among the transitional group of Rickettsia. | | The authors have provided a detailed explanation of this correction in the Supplementary File 1. | | The authors apologise for any inconvenience caused. |

Repeating the BinaxNOW antigen test for SARS-CoV-2 by two groups of readers within 30 minutes resulted in high concordance (98.9%) in 2,110 encounters. BinaxNOW test sensitivity was 77.2% (258/334) compared to real-time reverse transcription-polymerase chain reaction. Repeating antigen testing on the same day did not significantly improve test sensitivity while specificity remained high.Competing Interest StatementThe authors have declared no competing interest.Funding StatementThis work was funded by the Centers for Disease Control and Prevention.Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:This activity was reviewed by CDC and was conducted consistent with applicable federal law and CDC policy. See e.g., 45 C.F.R. part 46.102(l)(2), 21 C.F.R. part 56; 42 U.S.C. 241(d); 5 U.S.C. 552a; 44 U.S.C. 3501 et seq.All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).YesI have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable.YesData will be made available upon reasonable request.

The western black-legged tick (Ixodes pacificus) is the most frequently identified human-biting tick species in the western United States and the principal vector of at least three recognized bacterial pathogens of humans. A potentially pathogenic Rickettsia species, first described in 1978 and recently characterized as a novel transitional group agent designated as Rickettsia tillamookensis, also exists among populations of I. pacificus, although the distribution and frequency of this agent are poorly known. We evaluated DNA extracts from 348 host-seeking I. pacificus nymphs collected from 9 locations in five California counties, and from 916 I. pacificus adults collected from 24 locations in 13 counties, by using a real-time PCR designed specifically to detect DNA of R. tillamookensis. DNA of R. tillamookensis was detected in 10 (2.9%) nymphs (95% CI: 1.6-5.2%) and 17 (1.9%) adults (95% CI: 1.2-3.0%) from 11 counties of northern California. Although site-specific infection rates varied greatly, frequencies of infection remained consistently low when aggregated by stage, sex, habitat type, or geographical region. Four novel isolates of R. tillamookensis were cultivated in Vero E6 cells from individual adult ticks collected from Alameda, Nevada, and Yolo counties. Four historical isolates, serotyped previously as 'Tillamook-like' strains over 40 yr ago, were revived from long-term storage in liquid nitrogen and confirmed subsequently by molecular methods as isolates of R. tillamookensis. The potential public health impact of R. tillamookensis requires further investigation.

A unique genotype of Rickettsia parkeri, designated R. parkeri strain Black Gap, has thus far been associated exclusively with the North American tick, Dermacentor parumapertus. The compete genome consists of a single circular chromosome with 1,329,522 bp and a G+C content of 32.5%.

The tick-borne pathogen Rickettsia parkeri causes a mild rickettsiosis, with cases reported from several countries to its known distribution in the Americas. Molecular analyses have identified a clear distinction between strains of R. parkeri sensu stricto (s. s.) and R. parkeri sensu lato (s. l.) as well as separation between North American and South American R. parkeri s. s. strains. To expand on this previous work, we developed a multilocus sequence typing analysis with two aims: first, to investigate the genetic diversity within strains of North American R. parkeri s. s., and second, to further the understanding of the genetic relationships between R. parkeri s. s. and R. parkeri s. l. Sixty-four R. parkeri isolates and 12 R. parkeri-positive tick lysates were analyzed using a novel typing scheme consisting of four coding regions and two intergenic regions. A concatenated Bayesian phylogeny that identified eight clades was constructed: three represent the R. parkeri s. l. strains, and five represent the R. parkeri s. s. strains. The clades appear to be generally phylogeographically organized and associated with specific tick vectors. However, while one of the four R. parkeri s. s. North American clades appears to be limited to the southwestern United States, the other North American clades exhibit broad dispersal, most notably seen in the largest group, which includes representative samples extending from northern Mexico to Delaware. This work highlights the increasingly recognized geographic range of R. parkeri in the Americas and suggests a potential public health risk for these areas. IMPORTANCE Since 1937, when Rickettsia parkeri was originally identified in Amblyomma maculatum group ticks, the recognized range and associated vectors for this pathogen have expanded significantly. In recent years, R. parkeri has been identified in 12 tick species from seven countries in the Americas. Herein, we provide evidence that the greatest genetic diversity within R. parkeri exists in North America, where one R. parkeri sensu lato and four R. parkeri sensu stricto genotypes are present. While one distinct R. parkeri sensu stricto genotype exists only in the southwestern United States, three genotypes are broadly distributed in the eastern United States, with the largest of these found across the known range of R. parkeri in North America. In contrast, the South American R. parkeri sensu stricto samples represent a single genotype and are completely clonal at the loci analyzed, irrespective of their country of origin.

A previously unrecognized Rickettsia species was isolated in 1976 from a pool of Ixodes pacificus ticks collected in 1967 from Tillamook County, Oregon, USA. The isolate produced low fever and mild scrotal oedema following intraperitoneal injection into male guinea pigs (Cavia porcellus). Subsequent serotyping characterized this isolate as distinct from recognized typhus and spotted fever group Rickettsia species; nonetheless, the isolate remained unevaluated by molecular techniques and was not identified to species level for the subsequent 30 years. Ixodes pacificus is the most frequently identified human-biting tick in the western United States, and as such, formal identification and characterization of this potentially pathogenic Rickettsia species is warranted. Whole-genome sequencing of the Tillamook isolate revealed a genome 1.43 Mbp in size with 32.4 mol% G+C content. Maximum-likelihood phylogeny of core proteins places it in the transitional group of Rickettsia basal to both Rickettsia felis and Rickettsia asembonensis. It is distinct from existing named species, with maximum average nucleotide identity of 95.1% to R. asembonensis and maximum digital DNA-DNA hybridization score similarity to R. felis at 80.1%. The closest similarity at the 16S rRNA gene (97.9%) and sca4 (97.5%/97.6% respectively) is to Candidatus 'Rickettsia senegalensis' and Rickettsia sp. cf9, both isolated from cat fleas (Ctenocephalides felis). We characterized growth at various temperatures and in multiple cell lines. The Tillamook isolate grows aerobically in Vero E6, RF/6A and DH82 cells, and growth is rapid at 28 °C and 32 °C. Using accepted genomic criteria, we propose the name Rickettsia tillamookensis sp. nov., with the type strain Tillamook 23. Strain Tillamook 23 is available from the Centers for Disease Control and Prevention Rickettsial Isolate Reference Collection (WDCM 1093), Atlanta, GA, USA (CRIRC accession number RTI001(T)) and the Collection de Souches de l'Unité des Rickettsies (WDCM 875), Marseille, France (CSUR accession number R5043). Using accepted genomic criteria, we propose the name Rickettsia tillamookensis sp. nov., with the type strain Tillamook 23 (=CRIRC RTI001=R5043).

Repeating the BinaxNOW antigen test for SARS-CoV-2 by two groups of readers within 30 minutes resulted in high concordance (98.9%) in 2,110 encounters. BinaxNOW test sensitivity was 77.2% (258/334) compared to real-time reverse transcription-polymerase chain reaction. Same day antigen testing did not significantly improve test sensitivity while specificity remained high.

Ehrlichia chaffeensis causes human monocytic ehrlichiosis, and its principal vector is the Amblyomma americanum tick. The most frequently identified cases of ehrlichiosis come from the southeastern and south central states of the United States. In this study, a molecular typing system was developed that allows for the genetic differentiation of E. chaffeensis isolates. This multi-locus typing system included sequencing and analyzing intergenic regions ECH0033-ECH0035 and ECH0217-ECH0218, plus, variable genes variable length PCR target, 28-kDa, 120-kDa, and hemE. We examined a total of 31 unique isolates from humans and white-tailed deer, and eight DNA samples extracted from infected A. americanum collected from multiple states. This is the largest evaluation of E. chaffeensis isolates and their genotypes. Our findings show that when sequences of all six loci were concatenated and compared, the 39 samples could be separated into 23 genotypes and further grouped into six phylogenetic clades. The data in this study show no clear pattern between the geographic alignment with the genetic differentiation between the strains. As a result, this poses a challenge to understanding the spread of E. chaffeensis in the United States. Interestingly, our findings indicate that multiple strains from distant geographic origins share the same mutations, which suggests that the strains are being moved from one site to another by their hosts or vectors. In addition, we are seeing a northward shift in the lone star tick distribution in the United States. Last, some data also suggest minimal genetic mutations have occurred over time among strains that are within geographical proximity.

Genetic diversity of Anaplasma phagocytophilum was assessed in specimens from 16 infected patients and 16 infected Ixodes scapularis ticks. A region immediately downstream of the 16S rRNA gene, which included the gene encoding SdhC, was sequenced. For the A. phagocytophilum strains from patients no sequence differences were detected in this region. In contrast, significantly fewer ticks had a sequence encoding SdhC that was identical to that of the human strains (11/16 vs. 16/16, p = 0.04). This variation is consistent with the premise that not all A. phagocytophilum strains present in nature are able to cause clinical illness in humans. A strain referred to as A. phagocytophilumVariant-1 that is regarded as non-pathogenic for humans was previously described using a different typing method. Data from the current study suggest that both typing methods are identifying the same non-pathogenic strains.

Rickettsia rickettsii, the etiological agent of Rocky Mountain spotted fever (RMSF), a life-threatening tick-borne disease that affects humans and various animal species, has been recognized in medicine and science for more than 100 years. Isolate-dependent differences in virulence of R. rickettsii have been documented for many decades; nonetheless, the specific genetic and phenotypic factors responsible for these differences have not been characterized. Using in vivo and in vitro methods, we identified multiple phenotypic differences among six geographically distinct isolates of R. rickettsii, representing isolates from the United States, Costa Rica, and Brazil. Aggregate phenotypic data, derived from growth in Vero E6 cells and from clinical and pathological characteristics following infection of male guinea pigs (Cavia porcellus), allowed separation of these isolates into three categories: non-virulent (Iowa); mildly virulent (Sawtooth and Gila), and highly virulent (Sheila Smith(T), Costa Rica, and Taiaçu). Transcriptional profiles of 11 recognized or putative virulence factors confirmed the isolate-dependent differences between a mildly and a highly virulent isolate. These data corroborate previous qualitative assessments of strain virulence and suggest further that a critical and previously underappreciated balance between bacterial growth and host immune response could leverage strain pathogenicity. Also, this work provide insight into isolate-specific microbiological factors that contribute to the outcome of RMSF and confirms the hypothesis that distinct rickettsial isolates also differ phenotypically, which could influence the severity of disease in vertebrate hosts.

The Amblyomma maculatum Koch group of ixodid ticks consists of three species: A. maculatum, A. triste, and A. tigrinum. However, since Koch described this group in 1844, the systematics of its members has been the subject of ongoing debate. This is especially true of A. maculatum and A. triste; recent molecular analyses reveal insufficient genetic divergence to separate these as distinct species. Further confounding this issue is the discovery in 2014 of A. maculatum group ticks in southern Arizona (AZ), USA, that share morphological characteristics with both A. triste and A. maculatum. To biologically evaluate the identity of A. maculatum group ticks from southern Arizona, we analyzed the reproductive compatibility between specimens of A. maculatum group ticks collected from Georgia (GA), USA, and southern Arizona. Female ticks from both Arizona and Georgia were mated with males from both the Georgia and Arizona Amblyomma populations, creating two homologous and two heterologous F1 cohorts of ticks: GA ♀/GA ♂, AZ ♀/AZ ♂, GA ♀/AZ ♂, and AZ ♀/GA ♂. Each cohort was maintained separately into the F2 generation with F1 females mating only with F1 males from their same cohort. Survival and fecundity parameters were measured for all developmental stages. The observed survival parameters for heterologous cohorts were comparable to those of the homologous cohorts through the F1 generation. However, the F1 heterologous females produced F2 egg clutches that did not hatch, thus indicating that the Arizona and Georgia populations of A. maculatum group ticks tested here represent different biological species.

Rickettsia akari, an obligately intracellular bacterium, is the causative agent of the cosmopolitan urban disease rickettsialpox. R. akari is an atypical representative of spotted fever group rickettsiae (SFG) as it is associated with rodent mites rather than ticks or fleas; however, only limited information is available about the degree of genetic variability found among isolates of R. akari. We examined 13 isolates of R. akari from humans, rodents and mites in the USA, the former Soviet Union, and the former Yugoslavia made between 1946 and 2003 for diversity in their tandem repeat regions (TR) and intergenic regions (IGR). The 1.23 Mb genome of R. akari strain Hartford CWPP was analyzed using Tandem Repeat Finder software (http://tandem.bu.edu) and 374 different TRs were identified, with size variation from 1 to 483 bp and with TR copy numbers ranging between 21 and 1.9, respectively. No size polymorphisms were detected among the 11 TR regions examined from 5 open reading frames and 6 IGR. Eighteen non-TR IGR’s were amplified and sequenced for the same isolates comprising a total of 5.995 bp (0.49%) of the Hartford CWPP strain chromosome. Three single nucleotide polymorphism (SNP) sites were detected in two IGR’s which permitted separation of the five R. akari isolates from Ukraine SSR from the other eight isolates. In conclusion, this is the first study reporting genetic heterogeneity among R. akari isolates of different geographic origins. Further exploration of this genetic diversity is needed to understand better the geographic distribution of R. akari and the epidemiology of rickettsialpox. The potential of mites as hosts for other rickettsial agents also needs further investigation.

Tick-borne diseases (TBDs) continue to emerge and re-emerge in several regions of the world, highlighting the need for novel and effective control strategies. The development of effective strategies requires a better understanding of TBDs ecology, and given the complexity of these systems, interdisciplinary approaches are required. In recent years, the microbiome of vectors has received much attention, mainly because associations between native microbes and pathogens may provide a new promising path towards the disruption of pathogen transmission. However, we still do not fully understand how host genetics and environmental factors interact to shape the microbiome of organisms, or how pathogenic microorganisms affect the microbiome and vice versa. The integration of different lines of evidence may be the key to improve our understanding of TBDs ecology. In that context, we generated microbiome and pathogen presence data for Dermacentor variabilis, and integrated those data sets with population genetic data, and metadata for the same individual tick specimens. Clustering and multivariate statistical methods were used to combine, analyze, and visualize data sets. Interpretation of the results is challenging, likely due to the low levels of genetic diversity and the high abundance of a few taxa in the microbiome. Francisella was dominant in almost all ticks, regardless of geography or sex. Nevertheless, our results showed that, overall, ticks from different geographic regions differ in their microbiome composition. Additionally, DNA of Rickettsia rhipicephali, R. montanensis, R. bellii, and Anaplasma spp., was detected in D. variabilis specimens. This is the first study that successfully generated microbiome, population genetics, and pathogen presence data from the same individual ticks, and that attempted to combine the different lines of evidence. The approaches and pre-processing steps used can be applied to a variety of taxa, and help better understand ecological processes in biological systems.

Amblyomma maculatum Koch sensu lato (s.l.) ticks are the vector of Rickettsia parkeri in Arizona, where nine cases of R. parkeri rickettsiosis have been identified since the initial case in 2014. The current study sought to better define the geographic ranges of the vector and pathogen and to assess the potential public health risk posed by R. parkeri in this region of the southwestern United States. A total of 275 A. maculatum s.l. ticks were collected from 34 locations in four counties in Arizona and one county in New Mexico and screened for DNA of Rickettsia species. Rickettsia parkeri was detected in 20.4% of the ticks, including one specimen collected from New Mexico, the first report of R. parkeri in A. maculatum s.l. from this state. This work demonstrates a broader distribution of A. maculatum s.l. ticks and R. parkeri in the southwestern United States than appreciated previously to suggest that R. parkeri rickettsiosis is underrecognized in this region.

Congregate work and residential locations are at increased risk for infectious disease transmission including respiratory illness outbreaks. SARS-CoV-2, the virus that causes coronavirus disease 2019 (COVID-19), is primarily spread person to person through respiratory droplets. Nationwide, the meat and poultry processing industry, an essential component of the U.S. food infrastructure, employs approximately 500,000 persons, many of whom work in proximity to other workers (1). Because of reports of initial cases of COVID-19, in some meat processing facilities, states were asked to provide aggregated data concerning the number of meat and poultry processing facilities affected by COVID-19 and the number of workers with COVID-19 in these facilities, including COVID-19-related deaths. Qualitative data gathered by CDC during on-site and remote assessments were analyzed and summarized. During April 9-27, aggregate data on COVID-19 cases among 115 meat or poultry processing facilities in 19 states were reported to CDC. Among these facilities, COVID-19 was diagnosed in 4,913 (approximately 3%) workers, and 20 COVID-19-related deaths were reported. Facility barriers to effective prevention and control of COVID-19 included difficulty distancing workers at least 6 feet (2 meters) from one another (2) and in implementing COVID-19-specific disinfection guidelines.* Among workers, socioeconomic challenges might contribute to working while feeling ill, particularly if there are management practices such as bonuses that incentivize attendance. Methods to decrease transmission within the facility include worker symptom screening programs, policies to discourage working while experiencing symptoms compatible with COVID-19, and social distancing by workers. Source control measures (e.g., the use of cloth face covers) as well as increased disinfection of high-touch surfaces are also important means of preventing SARS-CoV-2 exposure. Mitigation efforts to reduce transmission in the community should also be considered. Many of these measures might also reduce asymptomatic and presymptomatic transmission (3). Implementation of these public health strategies will help protect workers from COVID-19 in this industry and assist in preserving the critical meat and poultry production infrastructure (4).

Dermacentor andersoni, the Rocky Mountain wood tick, occurs predominantly in the northwestern United States and southwestern Canada. There are relatively few contemporary data to evaluate the occurrence of Rickettsia and Anaplasma species in D. andersoni in western North America, and even less information about these associations in the state of Washington, where this tick species is widely distributed and often bites humans. We used PCR assays to detect DNA of Rickettsia and Anaplasmataceae bacteria in 203 adult D. andersoni ticks collected from 17 sites in 9 counties of Washington between May 2012 and May 2015. Of these, 56 (27.6 %) were infected with a Rickettsia species and 3 (5.4 %) with a member of the Anaplasmataceae family. Rickettsia peacockii, R. bellii and R. rhipicephali were found in 17.7 %, 4.9 %, and 4.4 % of the Rickettsia positive ticks, respectively. Coinfections of R. bellii with R. peacockii or R. rhipicephali were identified in 6 ticks. Of the Anaplasmataceae-positive ticks, one was identified as being infected with Anaplasma phagocytophilum AP-Variant 1. No ticks were infected with a recognized human or animal pathogen, including R. rickettsii, A. phagocytophilum-ha, A. bovis, or A. marginale.

Rickettsia parkeri, a tick-borne pathogen distributed throughout several countries of the Americas, causes a mild to moderately severe, eschar-associated spotted fever rickettsiosis. Although most U.S. cases of R. parkeri rickettsiosis are reported from southeastern states, some have been reported recently from remote regions of southern Arizona. These cases are linked to R. parkeri-infected ticks of the Amblyomma maculatum (Acari: Ixodidae) group found in several isolated mountain ranges of southern Arizona and New Mexico, referred to as 'sky islands'. Archival records also document ticks of the A. maculatum group collected from domestic and wild animals in West Texas. We surveyed sites in two sky island chains of Jeff Davis and Brewster counties to document the off-host occurrence of these ticks and identify the presence of R. parkeri in the Trans-Pecos region of Texas. During August 2019, 43 adult A. maculatum group ticks were flagged from vegetation or removed from a road-killed, female mule deer. Of 39 samples evaluated by PCR, eight contained a partial sca0 sequence with complete identity to R. parkeri and two with complete identity to 'Candidatus Rickettsia andeanae', a species of undetermined pathogenicity. Four isolates of R. parkeri were obtained using cell culture. Persons at risk for R. parkeri rickettsiosis include those who work or recreate in these mountains, such as hikers, backpackers, research scientists, foresters, and border enforcement personnel. Additional investigations are needed to define the distribution of these medically important arthropods in other parts of the southwestern United States and northern Mexico.

We found Rickettsia parkeri in Amblyomma ovale ticks collected in Veracruz, Mexico, in 2018. We sequenced gene segments of gltA, htrA, sca0, and sca5; phylogenetic reconstruction revealed near-complete identity with R. parkeri strain Atlantic Rainforest. Enhanced surveillance is needed in Mexico to determine the public health relevance of this bacterium.

Pacific Coast tick fever is a febrile illness associated with the bite of Dermacentor occidentalis and results from an infection due to the intracellular pathogen Rickettsia 364D (also known by the proposed name "Rickettsia philipii"). Current molecular methods for the detection of this pathogen rely on the amplification of a conserved spotted fever group rickettsial gene (ompA) followed by DNA sequencing of the amplicon to identify the species. This work describes the development of a Rickettsia 364D-specific TaqMan assay to simplify and accelerate the detection and identification processes. The assay demonstrated a sensitivity of 1 genomic copy per 4 mul sample and is highly specific for Rickettsia 364D. The utility of this assay for ecological and diagnostic samples was evaluated using single-blinded banked specimens and yielded a clinical sensitivity and specificity of 100%. In conclusion, we describe the development and evaluation of a novel TaqMan real-time PCR assay for the detection and identification of Rickettsia 364D suitable for ecological and diagnostic applications.

Introduction of ticks into the United States that can carry disease-causing pathogens to humans, companion animals, and wildlife has accelerated in recent years, mostly due to globalization, frequency of travel, and a rise in legal and illegal animal trades. We hereby report for the first time introduction of a live fully engorged Amblyomma coelebs feeding on a human into the United States from Central America. Amblyomma coelebs is geographically distributed in the Neotropical region and reaches the southern states of Mexico. This species is capable of transmitting a number of pathogens of public health and veterinary importance including spotted fever group rickettsiae, raising concern that A. coelebs, if it became established in the United States, might also be able to carry these pathogens. Considering the risks of exotic ticks as vectors of numerous pathogens and their potential to establish new populations under conducive climatic and habitat conditions, rigorous inspection practices of imported livestock and pet animals at ports of entry are vital. It is also important for travelers and practitioners to develop a heightened awareness of the public health risks associated with the unintended importation of exotic ticks and the potential such parasites have for breaching United States biosecurity defenses.

We report Rickettsia parkeri and Candidatus Rickettsia andeanae in ticks of the Amblyomma maculatum group collected from dogs in Sonora, Mexico. Molecular characterization of these bacteria was accomplished by DNA amplification and sequence analysis of portions of the rickettsial genes gltA, htrA, ompA, and ompB.

Rickettsia parkeri is a recently recognized human pathogen transmitted in the southeastern United States by Amblyomma maculatum, the Gulf Coast tick. Since R. parkeri was conclusively identified as a human pathogen in 2004, over 40 cases of R. parkeri rickettsiosis have been identified in the United States, most of which occur in the southeastern states. During 2012-2014, five of these cases were identified by a single urgent care practice in Coweta County, a Georgia county within the Atlanta metropolitan area. To investigate the occurrence of R. parkeri-infected A. maculatum in the Atlanta metropolitan area, ticks were collected from 6 counties around the city of Atlanta and evaluated for infection with a Rickettsia species. A total of 263 questing adult A. maculatum were collected during 2015 and 2016. Of these, 93 (35%) were PCR-positive for DNA of R. parkeri and an additional 46 (17%) were PCR-positive for DNA of "Candidatus Rickettsia andeanae," a spotted fever group Rickettsia species of unknown pathogenicity. No co-infections of these two rickettsiae were detected; however four of the six counties sampled showed presence of both rickettsial organisms. The high frequency of R. parkeri in these tick populations indicates a potential risk for those living, working, or recreating in A. maculatum-infested habitats within these six counties in the Atlanta metropolitan area.

Dermacentor variabilis, a common human-biting tick found throughout the eastern half and along the west coast of the United States, is a vector of multiple bacterial pathogens. Historically, D. variabilis has been considered a primary vector of Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever. A total of 883 adult D. variabilis, collected between 2012 and 2017 from various locations in 12 states across the United States, were screened for rickettsial DNA. Tick extracts were evaluated using three real-time PCR assays; an R. rickettsii-specific assay, a Rickettsia bellii-specific assay, and a Rickettsia genus-specific assay. Sequencing of ompA gene amplicons generated using a seminested PCR assay was used to determine the rickettsial species present in positive samples not already identified by species-specific real-time assays. A total of 87 (9.9%) tick extracts contained R. bellii DNA and 203 (23%) contained DNA of other rickettsial species, including 47 (5.3%) with Rickettsia montanensis, 11 (1.2%) with Rickettsia amblyommatis, 2 (0.2%) with Rickettsia rhipicephali, and 3 (0.3%) with Rickettsia parkeri. Only 1 (0.1%) tick extract contained DNA of R. rickettsii. These data support multiple other contemporary studies that indicate infrequent detection of R. rickettsii in D. variabilis in North America.

Nosocomial infections of Elizabethkingia species can have fatal outcomes if not identified and treated properly. The current diagnostic tools available require culture and isolation, which can extend the reporting time and delay treatment. Using comparative genomics, we developed an efficient multiplex real-time PCR for the simultaneous detection of all known species of Elizabethkingia, as well as differentiating the two most commonly reported species Elizabethkingia anophelis and Elizabethkingia meningoseptica.

In the United States, the lone star tick, Amblyomma americanum (L.) (Acari: Ixodidae), is an aggressive southeastern species whose range has reportedly been steadily expanding northward. The number of A. americanum specimens submitted to the Tick Testing Laboratory (TTL) at the Connecticut Agricultural Experiment Station (CAES) increased by 58% from the period of 1996-2006 (n = 488) to 2007-2017 (n = 773), mainly from Fairfield County in the southwestern corner of the state. The greatest numbers of A. americanum submissions to the CAES-TTL were from the City of Norwalk and a few adjacent municipalities. We also report the discovery of a large infestation of adult and nymphal lone star ticks detected on a dead male white-tailed deer, Odocoileus virginianus (Zimmerman) (Artiodactyla: Cervidae), on Manresa Island, Norwalk, in June 2017, indicating a long established, undetected population along the southwestern coast. A sample of nymphal and adult host-seeking A. americanum collected July 2017 from Manresa Island were tested and a proportion were positive for Ehrlichia chaffeensis, Ehrlichia ewingii, and Anaplasma phagocytophilum. The A. americanum tick and its associated disease pathogens are expected to become an increasing public health concern in southern New England.

In the north-central United States, the blacklegged tick (Ixodes scapularis) is currently known to vector seven human pathogens. These include five bacteria (Borrelia burgdorferi sensu stricto, Borrelia mayonii, Borrelia miyamotoi, Anaplasma phagocytophilum, Ehrlichia muris eauclairensis), one protozoan (Babesia microti) and one virus (Powassan). We sought to assess the prevalence and distribution of these pathogens in host-seeking nymphs collected throughout Minnesota, a state on the northwestern edge of the tick's expanding range, where reported cases of I. scapularis-borne diseases have increased in incidence and geographic range over the past decade. Among the 1240 host-seeking I. scapularis nymphs that we screened from 64 sites, we detected all seven pathogens at varying frequencies. Borrelia burgdorferi s.s. was the most prevalent and geographically widespread, found in 25.24% of all nymphs tested. Anaplasma phagocytophilum and Babesia microti were also geographically widespread, but they were less prevalent than Bo. burgdorferi s.s. (detected in 6.29% and 4.68% of ticks, respectively). Spatial clusters of sites with high prevalence for these three pathogens were identified in the north-central region of the state. Prevalence was less than 1.29% for each of the remaining pathogens. Two or more pathogens were detected in 90 nymphs (7.26%); coinfections with Bo. burgdorferi s.s. and either A. phagocytophilum (51 nymphs, 4.11%) or Ba. microti (43 nymphs, 3.47%) were the most common combinations. The distribution and density of infected ticks mirrors the distribution of notifiable tick-borne diseases in Minnesota and provides information on the distribution and prevalence of recently described human pathogens.