Prevalence and Strains of Colorado Tick Fever Virus in Rocky Mountain Wood Ticks in the Bitterroot Valley, Montana.

The Rocky Mountain wood tick, Dermacentor andersoni, has long been known to transmit human pathogens. Within the Bitterroot Valley, Ravalli County, Montana, these agents include Rickettsia rickettsii, Francisella tularensis, and Colorado tick fever virus (CTFV). Found in the western United States where wood ticks occur, CTFV causes a biphasic, febrile illness in humans and persists in enzootic cycles involving the ticks and small mammals. CTFV belongs to the genus Coltivirus, family Reoviridae, whose genome consists of 12 double-stranded RNA segments. Previous studies revealed the presence of CTFV-infected ticks and rodents in select locations within the valley in the 1960s and 1970s, using animal and cell culture methods for detection. We aimed to determine the range and prevalence of the virus in adult questing ticks throughout the valley using molecular tools and to examine the genomic variation between virus strains. Adult D. andersoni ticks were collected during 2002-2003 and 2009-2013. RNA extractions and reverse transcription-polymerase chain reaction were performed on 921 ticks, of which 61 ticks were positive for CTFV, resulting in a 6.6% prevalence of infection. Four genetic loci, one from each of the segments 9, 10, 11, and 12, within the viral genome were sequenced. Reassortment was detected between CTFV sequence strains within the valley. This study confirmed the prevalence of CTFV in D. andersoni ticks within the Bitterroot Valley, which has remained at levels found in the 1950s and 60s. Additional CTFV sequences were obtained and evidence of reassortment was observed between strains within the valley.

Case Report: A Case of Colorado Tick Fever Acquired in Southwestern Saskatchewan.

Colorado tick fever virus is transmitted by Dermacentor andersoni ticks. In Canada, these ticks are found in the southern regions of British Columbia (Rocky Mountains) and Alberta, as well as southwestern Saskatchewan. Colorado tick fever should be clinically suspected in patients presenting with a biphasic febrile illness and leukopenia following tick exposure in the appropriate geographic area.

Detection of Colorado Tick Fever viral RNA in acute human serum samples by a quantitative real-time RT-PCR assay.

A quantitative real-time RT-PCR assay for the detection of Colorado Tick Fever (CTF) viral RNA in human clinical samples is presented. The sensitivity of this assay has been shown to be greater than that of the isolation of virus in Vero cells by standard plaque assay in a direct comparison. The specificity of the CTF quantitative real-time RT-PCR assay was determined by the exclusive detection of CTF viral RNAs when applied to a diverse panel of CTF viral isolates and reference strain agents known to circulate in areas of CTF virus transmission. Lastly, the quantitative real-time RT-PCR assay demonstrated exceptional sensitivity for the detection of CTF viral RNA in acute human serum. The quantitative real-time RT-PCR assay is efficient, sensitive and specific and as such is useful for the detection of CTF viral RNA in the diagnostic or research laboratory.

Colorado tick fever in the United States, 2002-2012.

BACKGROUND: Colorado tick fever (CTF) is an acute systemic febrile illness caused by the CTF virus (CTFV). The last national summary of CTF cases in the United States included cases reported through 2001. This study summarizes national surveillance data for CTF from 2002 through 2012 and examines trends in the epidemiology and testing of identified CTF cases. METHODS: Because CTF is not nationally notifiable, we identified CTF cases through solicited reports from state health departments and diagnostic laboratory records. For all cases, we collected data on age, sex, county of residence, travel history, symptom onset date, laboratory testing, and clinical outcome. Poisson regression was used to examine trends over time in case counts, and simple linear regression and logistic regression were used to examine trends in case characteristics. RESULTS: From 2002 through 2012, 75 CTF cases were identified with a median of five cases per year (range 3-14). Forty-seven (63%) cases occurred in males and 49 (65%) occurred in people aged ≥40 years. The majority (80%) of cases had onset of illness during May through July. Cases occurred in residents of 14 states but the infections were acquired in six western states. Wyoming had the highest annual incidence of CTF among residents (3.4 cases per million population), followed by Montana (1.5 per million), and Utah (0.5 per million). Over the 11 years, there was an increase in the proportion of cases diagnosed by RT-PCR testing and in the proportion of cases among travelers to another state. CONCLUSIONS: CTF cases continue to occur annually among residents and visitors to the western United States. Public health prevention messages about decreasing tick exposure should be targeted to residents and travelers who will spend time outdoors in an endemic region during the spring and summer months.

Experimental Colorado tick fever virus infection in Colorado mammals.

One hundred and twenty-four small mammals of six species were inoculated with three strains of Colorado tick fever (CTF) virus to define viremia and neutralizing (N) antibody responses. Adult Eutamias minimus and Eutamias embrinus, and juvenile Peromyscus maniculatus and Spermophilus lateralis, were highly susceptible to development of viremic infection. Adult S. lateralis and P. maniculatus were moderately susceptible (greater than or equal to 50% viremic). Five Sylvilagus nuttalli did not become viremic following experimental inoculation. Spermophilus richardsoni was also relatively resistant (less than or equal to 50% viremic). The longest duration of viremia (mean 15.8 days) and highest peak viremia levels (mean peak titer 10(3.9 plaque-forming units per ml) occurred in E. minimus. Adult E. umbrinus, juvenile S. lateralis, and juvenile P. maniculatus had moderate viremias. Adult S. lateralis and S. richardsoni often had short viremias during which virus was only intermittently detectable. N antibody production was most rapid in E. minimus in comparison with other species. In addition, N antibody persisted for 1 year in this species. In other species, many animals lost detectable antibody 5-11 months after infection. No significant differences were found in patterns of infection between three CTF virus strains. We conclude that of the six species inoculated, E. minimus is the best experimental host for CTF virus.

The ecology of Colorado tick fever in Rocky Mountain National Park in 1974. II. Infection in small mammals.

Field studies of Colorado tick fever (CTF) in small mammals in Rocky Mountain National Park (RMNP) in 1974 established that Eutamias minimus and Spermophilus lateralis were the most important hosts for CTF virus and were the source of virus for immature stages of the tick vector, Dermacentor andersoni. Other species (Peromyscus maniculatus, Spermophilus richardsonii, Eutamias umbrinus) are secondary hosts. The intensity of viral activity in rodents varied greatly from locality to locality. Highest rodent infection rates were found to occur in the Moraine Park area of RMNP. Lowest infection rates occurred above 3,290 meters in altitude at Rainbow Curve and on the tundra. The prevalence of infection in rodents was constant from April--July (5--6% of animals captured were viremic) and then declined to 1.7--2.5% in August and September coincident with a decline in nymphal tick ectoparasitism. Many animals were captured which were simultaneously viremic and antibody-positive. Under field conditions, neutralizing antibody seroconversion does not always occur.

Co-circulation of multiple Colorado tick fever virus genotypes.

Colorado tick fever (CTF) virus, family Reoviridae, genus Orbivirus, contains 12 genes distinguishable by polyacrylamide gel electrophoresis (PAGE). Multiple genotypes of CTF virus were isolated at 3 field sites in Colorado in 1985. Five genotypes were found at Campos Cabin, 2 at Drake, and 6 at Rocky Mountain National Park. Virus isolations were made in 1985 from 6 patients with CTF. These isolates were distinct from each other and the field isolates. Although the CTF isolates were different by PAGE profile, the majority of the 12 genes were highly conserved among the 1985 isolates and a Florio isolate (FMA). Only genes 4 and 6 were variant among the 1985 CTF isolates and FMA, and no unique genes were identified. In 1986, a follow-up field survey was done at the Campos Cabin site. Of the 3 CTF PAGE genotypes obtained, 2 exhibited PAGE profiles which were different from the 1985 isolates. One isolate may have resulted from the reassortment of genes from 2 of the isolates circulating at Campos Cabin in 1985.

The ecology of Colorado tick fever in Rocky Mountain National Park in 1974. III. Habitats supporting the virus.

Ecologic studies of small mammals in Rocky Mountain National Park (RMNP) were conducted in 1974 in order to identify the specific habitats within the Lower Montane Forest that support Colorado tick fever (CTF) virus. Data was collected on the abundance and distribution of 4 primary rodent species, tick infestation, CTF virus, and neutralizing antibody prevalence. Rodents were captured along transects crossing different habitats. Open stands of ponderosa pine and shrubs on dry, rocky surfaces were found to be important for maintaining CTF virus.

Survey for evidence of Colorado tick fever virus outside of the known endemic area in California.

A virus very similar or identical to Colorado tick fever (CTF) virus was recovered from the blood clot of one of 104 black-tailed jack rabbits (Lepus californicus) examined during a survey for various zoonotic agents in mammals and ticks from the University of California, Hopland Field Station, Mendocino County, California, 1974--79. This is the first reported isolation of a CTF-like virus from L. californicus, and only the second time such a virus has been found in northwestern California. Mendocino County is located far outside the known distributional ranges of the most common mammalian hosts of CTF virus and of Dermacentor andersoni, the only proven tick vector for man. The viral isolate is very similar to a CTF-like virus previously recovered from the blood and spleen of a western gray squirrel (Sciurus griseus) from San Luis Obispo County, an area also outside of the previously-known CTF area. Virus was not isolated from 14 additional species of mammals (354 specimens) or from eight species of ticks (4,487 individuals), but CTF-neutralizing antibodies were detected in 28 of 771 (3.6%) sera from seven of 15 mammalian species including significant titers (greater than or equal to 1:8) in two species and one subspecies not previously reported as natural hosts, i.e., brush mouse (Peromyscus boylii), pinyon mouse (P. truei), and Columbian black-tailed deer (Odocoileus hemionus columbianus). CTF indirect immunofluorescent antibodies also were detected in 26 of 129 (20.2%) sera belonging to four of five mammalian species tested. Neutralizing antibodies were found in sera of deer from other localities in Mendocino County, from a deer mouse from Napa County, and from a brush rabbit from Monterey County as well. These findings suggest that a virus identical or similar to CTF virus is widespread in northwestern-westcentral California, and that surveillance for human cases of CTF or a similar disease should be extended to cover this region.

Discriminant analysis: a method of identifying foci of vector-borne diseases.

Identification of foci of vector-borne diseases does not require knowledge of exact abundances of vectors and pathogens; rather, a focus is defined by the presence, or some minimal level of abundance, of a vector and pathogen. Stepwise discriminant analysis (DA) was applied to data on free-ranging adult wood ticks (the vector) and to data on isolations of Colorado tick fever virus from small mammals. Trap stations were grouped on the basis of relative abundance of wood ticks for one set of analyses and on the presence or absence of virus for another set of analyses. Additional data consisted of easily measured environmental variables. It is concluded that DA provides a useful tool for analysis of ecosystem structure and an effective means of identifying foci of infection.

Serologic and molecular diagnosis of Colorado tick fever viral infections.

Molecular and serologic methods usable for the biological diagnosis of Coltivirus infection are reported. We designed a multiplex reverse transcription-polymerase chain reaction system that allowed the simultaneous and specific amplification of three genomic segments from as little as 0.01 plaque-forming units. Another system in the S2 viral segment permitted the differential diagnosis of American and European viral isolates. We also discuss some improvements of previous ELISAs, and the results obtained with paired sera from Colorado tick fever (CTF) virus-infected individuals. Western blot analysis was developed that allowed the detection of antibodies to a 38-kD viral protein in all tested sera. It also enabled the detection of anti-CTF virus antibodies in ELISA-negative sera. Specific IgM antibodies against a synthetic viral peptide could be detected in sera at the acute stage of the infection. Together, these results should permit the diagnosis of Coltivirus infection at any stage of the pathology.

Colorado tick fever.

Colorado tick fever, also known as mountain fever and mountain tick fever, is a well-described, viral, tick-borne disease common to the Rocky Mountain region of the United States and Canada. The Rocky Mountain wood tick, Dermacentor andersoni, is the primary vector. The triad of high fever, severe myalgia, and headache is typical, but not specific. Although a self-limited disease in most cases, severe complications may occur. PCR techniques have been developed that allow the diagnosis to be established from the first day of symptoms. Ribavirin may merit consideration in the appropriate clinical setting.

Tick-borne diseases in the United States: Rocky Mountain spotted fever and Colorado tick fever. A review.

The historical, clinical, ecological, and epidemiological features of Rocky Mountain spotted fever and Colorado tick fever, the two important tick-borne diseases in the United States, are reviewed. Rocky Mountain spotted fever, once considered a disease of the past, has again become a measurable public health problem. Its nationwide incidence has steadily increased since 1960 and has reached record proportions in 1976. The various factors responsible for this trend as well as for the mortality rates, which in spite of availability of effective antibiotics ranges from 5 to 10%, are discussed. Education of the public about ticks and their potential role as vectors of Rickettsia rickettsii and/or Colorado tick fever virus, and about the clinical manifestations of Rocky Mountain spotted fever, is considered the best means for preventing high incidence and mortality from these diseases.

Ecology of porcupines (Erethizon dorsatum) and Colorado tick fever virus in Rocky Mountain National Park, 1975-1977.

The involvement of porcupines, Erethizon dorsatum (L.), in the ecology of Colorado tick fever (CTF) virus in Rocky Mountain National Park was investigated from 1975 to 1977. Porcupine dens and feeding activity were found mostly on rocky knolls or on south-facing slopes within open stands of the montane coniferous forest, and 20 adult porcupines were trapped or captured by hand at those locations. An average of 24.6 +/- 5.4 adult Dermacentor andersoni Stiles ticks were found per animal (annual range, 17.5-31.4 ticks). The minimum CTF virus infection rate of the ticks removed from porcupines varied from 129 to 257, whereas for questing adult D. andersoni ticks from the same geographic area was 205 in 1976. No virus was isolated from the 20 animals, but 85% had neutralizing antibody against CTF virus. Porcupines utilize the same habitats described for the CTF ecosystem in Rocky Mountain National Park and appear to be an important host for adult D. andersoni.

Infection with Colorado tick fever virus among humans and ticks in a national park and forest, Wyoming, 2010.

BACKGROUND: Colorado tick fever (CTF) is an underreported tick-borne viral disease occurring in the western United States. CTF illness includes fever, headache, and severe myalgia lasting for weeks. Wyoming has one of the highest CTF incidence rates with approximately 30% of infected persons reporting tick exposure in a Wyoming National Park or Forest before symptom onset. We assessed CTF virus infections among humans and Dermacentor andersoni ticks in Grand Teton National Park (GRTE) and Bridger-Teton National Forest (BTNF). METHODS: In June of 2010, 526 eligible employees were approached to participate in a baseline and 3-month follow-up serosurvey and risk behavior survey. Seropositivity was defined as antibody titers against CTF virus ≥10, as measured by the plaque reduction neutralization test. Ticks were collected at 27 sites within GRTE/BTNF and tested by RT-PCR for the CTF virus. RESULTS: A total of 126 (24%) employees participated in the baseline and follow-up study visits. Three (2%) employees were seropositive for CTF virus infection at baseline. During the study, 47 (37%) participants found unattached ticks on themselves, and 12 (10%) found attached ticks; however, no participants seroconverted against CTF virus. Walking through sagebrush (p=0.04) and spending time at ≥7000 feet elevation (p<0.01) were significantly associated with tick exposure. Ninety-nine percent (174/176) of ticks were D. andersoni, and all were found at ≥7000 feet elevation in sagebrush areas; 37 (21%) ticks tested positive for CTF virus and were found at 10 (38%) of 26 sites sampled. CONCLUSIONS: Although no GRTE or BTNF employees were infected with CTF virus during the study period, high rates of infected ticks were identified in areas with sagebrush at ≥7000 feet. CTF education and personal protection measures against tick exposure should be targeted to visitors and employees traveling to the high-risk environs identified in this study.

Detection of Colorado tick fever virus by using reverse transcriptase PCR and application of the technique in laboratory diagnosis.

Colorado tick fever (CTF) virus elicits an acute illness in humans, producing nonspecific flu-like symptoms and a biphasic fever in approximately 50% of patients. The disease is transmitted by the adult Rocky Mountain wood tick (Dermacentor andersoni), and therefore incidence is limited by the habitat and life cycle of that vector. The early symptoms of infection are difficult to distinguish from those of several other agents, especially Rickettsia rickettsii. Serologic testing is usually unable to provide evidence of CTF viral infection during the acute phase because of the late appearance of the various antibodies. Here we report the development and clinical application of a test to diagnose this disease during the acute stages. Oligonucleotide primers to the S2 segment of CTF (Florio) virus were made, and these were used in the amplification of a 528-bp fragment of DNA, transcribed from the double-stranded CTF virus RNA template by reverse transcriptase PCR. RNAs processed from 16 CTF virus isolates yielded similar results when analyzed on agarose gels. These were distinguishable from their antigenic relatives Eyach, S6-14-03, and T5-2092 and from other coltiviruses and an orbivirus but not from the antigenically distinct CTF virus-related isolate 720896. A mouse model demonstrated the utility of this method with whole-blood specimens, and CTF virus was successfully detected in human sera from the initial day of the onset of symptoms to 8 days later. The reverse transcriptase PCR method is a promising tool for the early diagnosis of CTF viral infection, or for ruling out CTF virus as the etiologic agent, in order to facilitate appropriate medical support.

Antigenic variants of Colorado tick fever virus.

Twenty strains of Colorado tick fever (CTF) virus, isolated from ticks, mammals and humans, and two antigenic relatives of CTF virus were compared in cross-neutralization tests. Viruses were tested using single-inoculation sera prepared in hamsters. Antigenic variation, as measured by differences detected in the neutralization test, was noted among the virus isolates identified as strains of CTF virus. The virus strains isolated from humans appeared to vary the most in serological reactions. The two antigenic relatives of CTF virus are clearly distinct from strains of CTF and are different from each other. Antigenic relationships between these two viruses were established using two sets of single-inoculation antisera and both complement fixation and neutralization tests. Six distinct antigenic variants of CTF virus isolated from humans and the virus strain from ticks (75V1906) that showed the least antigenic variation, were tested against 49 coded serum pairs from clinically diagnosed cases of CTF. Significant differences were noted in the number of convalescent-phase sera that reacted with each virus strain and in the number of seroconversions observed with each test virus strain. Convalescent phase sera that reacted with multiple virus strains often varied significantly in antibody titre from one virus strain to another. This indicates that, in some instances, antibody was probably produced in response to infection by different antigenic variants of CTF virus.

Isolation of Eyach virus (Reoviridae, Colorado tick fever group) from Ixodes ricinus and I. ventalloi ticks in France.

During the spring of 1981, two strains of an identical virus were isolated in Mayenne, France, from Ixodes ricinus L. and Ixodes ventalloi Gil Collado, parasiting a wild rabbit (Oryctolagus cuniculus L.). Virological and serological results indicated that these strains belong to the Colorado tick fever serogroup and are indistinguishable from Eyach virus, a tick-borne virus previously described from West Germany. Study of dsRNA PAGE profiles confirmed these conclusions. The morphology and morphogenesis of French isolates were studied in infected suckling mouse brains showing some peculiar features previously described for Colorado tick fever virus. The importance of the isolation of Eyach virus in France is discussed.

Diagnosis of Colorado tick fever virus infection by enzyme immunoassays for immunoglobulin M and G antibodies.

An immunoglobulin M (IgM) capture enzyme immunoassay technique was adapted for the detection of antibody to Colorado tick fever virus in sera from 84 individuals for whom diagnosis had been confirmed by virus isolation or neutralization test. Titers were compared with those for IgG and neutralizing antibodies in these Colorado tick fever cases. IgM antibody titers were higher than neutralizing antibody titers, but neither appeared until 1 to 2 weeks after the onset of illness. Neutralizing antibodies were detected earlier than IgM antibodies, and both were detected with greater frequency than IgG antibodies. Late-convalescent-phase sera contained both neutralizing and IgG antibodies, but IgM was all but undetectable by 2 months after onset. Although the neutralization test may remain the serological test of choice, the enzyme immunoassay for IgM antibody offers a simple and more rapid method of serodiagnosis; the enzyme immunoassay is, however, less sensitive than the neutralization test. Furthermore, because there was a sharp decline in IgM antibody after 45 days, the presence of IgM antibody in a single serum sample provides a basis for the presumptive serodiagnosis of recent Colorado tick fever virus infection.