Genetic Tracking of a Rabid Coyote (Canis latrans) Detected beyond a Rabies Enzootic Area in West Virginia, US.

Wildlife translocation and cross-species transmission can impede control and elimination of emerging zoonotic diseases. Tracking the geographic origin of both host and virus (i.e., translocation versus local infection) may help determine the most effective response when high-risk cases of emerging pathogens are identified in wildlife. In May 2022, a coyote (Canis latrans) infected with the raccoon (Procyon lotor) rabies virus variant (RRV) was collected in Lewis County, West Virginia, US, an area free from RRV. We applied host population genomics and RRV phylogenetic analyses to determine the most likely geographic origin of the rabid coyote. Coyote genomic analyses included animals from multiple eastern states bordering West Virginia, with the probable origin of the rabid coyote being the county of collection. The RRV phylogenetic analyses included cases detected from West Virginia and neighboring states, with most similar RRV sequences collected in a county 80 km to the northeast, within the oral rabies vaccination zone. The combined results suggest that the coyote was infected in an RRV management area and carried the RRV to Lewis County, a pattern consistent with coyote local movement ecology. Distant cross-species transmission and subsequent host movement presents a low risk for onward transmission in raccoon populations. This information helped with emergency response decision-making, thereby saving time and resources.

Detection of Apparent Early Rabies Infection by LN34 Pan-Lyssavirus Real-Time RT-PCR Assay in Pennsylvania.

The Pennsylvania Department of Health Bureau of Laboratories (PABOL) tested 6855 animal samples for rabies using both the direct fluorescent antibody test (DFA) and LN34 pan-lyssavirus reverse transcriptase quantitative PCR (RT-qPCR) during 2017-2019. Only two samples (0.03%) were initially DFA negative but positive by LN34 RT-qPCR. Both cases were confirmed positive upon re-testing at PABOL and confirmatory testing at the Centers for Disease Control and Prevention by LN34 RT-qPCR and DFA. Rabies virus sequences from one sample were distinct from all positive samples processed at PABOL within two weeks, ruling out cross-contamination. Levels of rabies virus antigen and RNA were low in all brain structures tested, but were higher in brain stem and rostral spinal cord than in cerebellum, hippocampus or cortex. Taken together, the low level of rabies virus combined with higher abundance in more caudal brain structures suggest early infection. These cases highlight the increased sensitivity and ease of interpretation of LN34 RT-qPCR for low positive cases.

Illuminating uveitis: metagenomic deep sequencing identifies common and rare pathogens.

BACKGROUND: Ocular infections remain a major cause of blindness and morbidity worldwide. While prognosis is dependent on the timing and accuracy of diagnosis, the etiology remains elusive in ~50 % of presumed infectious uveitis cases. The objective of this study is to determine if unbiased metagenomic deep sequencing (MDS) can accurately detect pathogens in intraocular fluid samples of patients with uveitis. METHODS: This is a proof-of-concept study, in which intraocular fluid samples were obtained from five subjects with known diagnoses, and one subject with bilateral chronic uveitis without a known etiology. Samples were subjected to MDS, and results were compared with those from conventional diagnostic tests. Pathogens were identified using a rapid computational pipeline to analyze the non-host sequences obtained from MDS. RESULTS: Unbiased MDS of intraocular fluid produced results concordant with known diagnoses in subjects with (n = 4) and without (n = 1) uveitis. Samples positive for Cryptococcus neoformans, Toxoplasma gondii, and herpes simplex virus 1 as tested by a Clinical Laboratory Improvement Amendments-certified laboratory were correctly identified with MDS. Rubella virus was identified in one case of chronic bilateral idiopathic uveitis. The subject's strain was most closely related to a German rubella virus strain isolated in 1992, one year before he developed a fever and rash while living in Germany. The pattern and the number of viral identified mutations present in the patient's strain were consistent with long-term viral replication in the eye. CONCLUSIONS: MDS can identify fungi, parasites, and DNA and RNA viruses in minute volumes of intraocular fluid samples. The identification of chronic intraocular rubella virus infection highlights the eye's role as a long-term pathogen reservoir, which has implications for virus eradication and emerging global epidemics.

Laboratory Testing for Middle East Respiratory Syndrome Coronavirus, California, USA, 2013-2014.

Since Middle East respiratory syndrome coronavirus (MERS-CoV) first emerged, the California Department of Public Health has coordinated efforts to identify possible cases in travelers to California, USA, from affected areas. During 2013-2014, the department investigated 54 travelers for MERS-CoV; none tested positive, but 32 (62%) of 52 travelers with suspected MERS-CoV had other respiratory viruses.

Assessing prevention measures and Sin Nombre hantavirus seroprevalence among workers at Yosemite National Park.

BACKGROUND: During 2012, a total of 10 overnight visitors to Yosemite National Park (Yosemite) became infected with a hantavirus (Sin Nombre virus [SNV]); three died. SNV infections have been identified among persons with occupational exposure to deer mice (Peromyscus maniculatus). METHODS: We assessed SNV infection prevalence, work and living environments, mice exposures, and SNV prevention training, knowledge, and practices among workers of two major employers at Yosemite during September-October, 2012 by voluntary blood testing and a questionnaire. RESULTS: One of 526 participants had evidence of previous SNV infection. Participants reported frequently observing rodent infestations at work and home and not always following prescribed safety practices for tasks, including infestation cleanup. CONCLUSION: Although participants had multiple exposures to deer mice, we did not find evidence of widespread SNV infections. Nevertheless, employees working around deer mice should receive appropriate training and consistently follow prevention policies for high-risk activities.

A novel outbreak enterovirus D68 strain associated with acute flaccid myelitis cases in the USA (2012-14): a retrospective cohort study.

BACKGROUND: Enterovirus D68 was implicated in a widespread outbreak of severe respiratory illness across the USA in 2014 and has also been reported sporadically in patients with acute flaccid myelitis. We aimed to investigate the association between enterovirus D68 infection and acute flaccid myelitis during the 2014 enterovirus D68 respiratory outbreak in the USA. METHODS: Patients with acute flaccid myelitis who presented to two hospitals in Colorado and California, USA, between Nov 24, 2013, and Oct 11, 2014, were included in the study. Additional cases identified from Jan 1, 2012, to Oct 4, 2014, via statewide surveillance were provided by the California Department of Public Health. We investigated the cause of these cases by metagenomic next-generation sequencing, viral genome recovery, and enterovirus D68 phylogenetic analysis. We compared patients with acute flaccid myelitis who were positive for enterovirus D68 with those with acute flaccid myelitis but negative for enterovirus D68 using the two-tailed Fisher's exact test, two-sample unpaired t test, and Mann-Whitney U test. FINDINGS: 48 patients were included: 25 with acute flaccid myelitis, two with enterovirus-associated encephalitis, five with enterovirus-D68-associated upper respiratory illness, and 16 with aseptic meningitis or encephalitis who tested positive for enterovirus. Enterovirus D68 was detected in respiratory secretions from seven (64%) of 11 patients comprising two temporally and geographically linked acute flaccid myelitis clusters at the height of the 2014 outbreak, and from 12 (48%) of 25 patients with acute flaccid myelitis overall. Phylogenetic analysis revealed that all enterovirus D68 sequences associated with acute flaccid myelitis grouped into a clade B1 strain that emerged in 2010. Of six coding polymorphisms in the clade B1 enterovirus D68 polyprotein, five were present in neuropathogenic poliovirus or enterovirus D70, or both. One child with acute flaccid myelitis and a sibling with only upper respiratory illness were both infected by identical enterovirus D68 strains. Enterovirus D68 viraemia was identified in a child experiencing acute neurological progression of his paralytic illness. Deep metagenomic sequencing of cerebrospinal fluid from 14 patients with acute flaccid myelitis did not reveal evidence of an alternative infectious cause to enterovirus D68. INTERPRETATION: These findings strengthen the putative association between enterovirus D68 and acute flaccid myelitis and the contention that acute flaccid myelitis is a rare yet severe clinical manifestation of enterovirus D68 infection in susceptible hosts. FUNDING: National Institutes of Health, University of California, Abbott Laboratories, and the Centers for Disease Control and Prevention.

Use of clinical and neuroimaging characteristics to distinguish temporal lobe herpes simplex encephalitis from its mimics.

BACKGROUND: We describe the spectrum of etiologies associated with temporal lobe (TL) encephalitis and identify clinical and radiologic features that distinguish herpes simplex encephalitis (HSE) from its mimics. METHODS: We reviewed all adult cases of encephalitis with TL abnormalities on magnetic resonance imaging (MRI) from the California Encephalitis Project. We evaluated the association between specific clinical and MRI characteristics and HSE compared with other causes of TL encephalitis and used multivariate logistic modeling to identify radiologic predictors of HSE. RESULTS: Of 251 cases of TL encephalitis, 43% had an infectious etiology compared with 16% with a noninfectious etiology. Of infectious etiologies, herpes simplex virus was the most commonly identified agent (n = 60), followed by tuberculosis (n = 8) and varicella zoster virus (n = 7). Of noninfectious etiologies, more than half (n = 21) were due to autoimmune disease. Patients with HSE were older (56.8 vs 50.2 years; P = .012), more likely to be white (53% vs 35%; P = .013), more likely to present acutely (88% vs 64%; P = .001) and with a fever (80% vs 49%; P < .001), and less likely to present with a rash (2% vs 15%; P = .010). In a multivariate model, bilateral TL involvement (odds ratio [OR], 0.38; 95% confidence interval [CI], .18-.79; P = .010) and lesions outside the TL, insula, or cingulate (OR, 0.37; 95% CI, .18-.74; P = .005) were associated with lower odds of HSE. CONCLUSIONS: In addition to HSE, other infectious and noninfectious etiologies should be considered in the differential diagnosis for TL encephalitis, depending on the presentation. Specific clinical and imaging features may aid in distinguishing HSE from non-HSE causes of TL encephalitis.

Measles outbreak--California, December 2014-February 2015.

On January 5, 2015, the California Department of Public Health (CDPH) was notified about a suspected measles case. The patient was a hospitalized, unvaccinated child, aged 11 years with rash onset on December 28. The only notable travel history during the exposure period was a visit to one of two adjacent Disney theme parks located in Orange County, California. On the same day, CDPH received reports of four additional suspected measles cases in California residents and two in Utah residents, all of whom reported visiting one or both Disney theme parks during December 17-20. By January 7,seven California measles cases had been confirmed, and CDPH issued a press release and an Epidemic Information Exchange (Epi-X) notification to other states regarding this outbreak. Measles transmission is ongoing.

Acute Flaccid Myelitis of Unknown Etiology in California, 2012-2015.

IMPORTANCE: There has been limited surveillance for acute flaccid paralysis in North America since the regional eradication of poliovirus. In 2012, the California Department of Public Health received several reports of acute flaccid paralysis cases of unknown etiology. OBJECTIVE: To quantify disease incidence and identify potential etiologies of acute flaccid paralysis cases with evidence of spinal motor neuron injury. DESIGN, SETTING, AND PARTICIPANTS: Case series of acute flaccid paralysis in patients with radiological or neurophysiological findings suggestive of spinal motor neuron involvement reported to the California Department of Public Health with symptom onset between June 2012 and July 2015. Patients meeting diagnostic criteria for other acute flaccid paralysis etiologies were excluded. Cerebrospinal fluid, serum samples, nasopharyngeal swab specimens, and stool specimens were submitted to the state laboratory for infectious agent testing. MAIN OUTCOMES AND MEASURES: Case incidence and infectious agent association. RESULTS: Fifty-nine cases were identified. Median age was 9 years (interquartile range [IQR], 4-14 years; 50 of the cases were younger than 21 years). Symptoms that preceded or were concurrent included respiratory or gastrointestinal illness (n = 54), fever (n = 47), and limb myalgia (n = 41). Fifty-six patients had T2 hyperintensity of spinal gray matter on magnetic resonance imaging and 43 patients had cerebrospinal fluid pleocytosis. During the course of the initial hospitalization, 42 patients received intravenous steroids; 43, intravenous immunoglobulin; and 13, plasma exchange; or a combination of these treatments. Among 45 patients with follow-up data, 38 had persistent weakness at a median follow-up of 9 months (IQR, 3-12 months). Two patients, both immunocompromised adults, died within 60 days of symptom onset. Enteroviruses were the most frequently detected pathogen in either nasopharynx swab specimens, stool specimens, serum samples (15 of 45 patients tested). No pathogens were isolated from the cerebrospinal fluid. The incidence of reported cases was significantly higher during a national enterovirus D68 outbreak occurring from August 2014 through January 2015 (0.16 cases per 100,000 person-years) compared with other monitoring periods (0.028 cases per 100,000 person-years; P <.001). CONCLUSIONS AND RELEVANCE: In this series of patients identified in California from June 2012 through July 2015, clinical manifestations indicated a rare but distinct syndrome of acute flaccid paralysis with evidence of spinal motor neuron involvement. The etiology remains undetermined, most patients were children and young adults, and motor weakness was prolonged.

Notes from the field: measles - California, January 1-April 18, 2014.

Measles is a highly contagious, acute viral illness that can lead to severe complications and death. Even patients who experience uncomplicated acute measles have a small risk for developing a devastating neurologic illness, subacute sclerosing panencephalitis, years after their infection. Measles was documented as eliminated (defined as interruption of continuous transmission lasting ≥12 months) in the United States in 2000; however, importation of measles cases and limited local transmission continue to occur. During January 1-April 18, 2014, the California Department of Public Health received reports of 58 confirmed measles cases, the highest number reported for that period since 1995. Patients ranged in age from 5 months to 60 years. Three (5%) patients were aged <12 months, six (10%) were aged 1-4 years, 17 (29%) were aged 5-19 years, and 32 (55%) were aged ≥20 years. As of April 18, there had been 12 hospitalizations, and no deaths had been reported. During 2000-2013, the median annual number of measles cases reported in California was nine (range = four to 40).

Hantavirus infections among overnight visitors to Yosemite National Park, California, USA, 2012.

In summer 2012, an outbreak of hantavirus infections occurred among overnight visitors to Yosemite National Park in California, USA. An investigation encompassing clinical, epidemiologic, laboratory, and environmental factors identified 10 cases among residents of 3 states. Eight case-patients experienced hantavirus pulmonary syndrome, of whom 5 required intensive care with ventilatory support and 3 died. Staying overnight in a signature tent cabin (9 case-patients) was significantly associated with becoming infected with hantavirus (p<0.001). Rodent nests and tunnels were observed in the foam insulation of the cabin walls. Rodent trapping in the implicated area resulted in high trap success rate (51%), and antibodies reactive to Sin Nombre virus were detected in 10 (14%) of 73 captured deer mice. All signature tent cabins were closed and subsequently dismantled. Continuous public awareness and rodent control and exclusion are key measures in minimizing the risk for hantavirus infection in areas inhabited by deer mice.

Five-year surveillance of West Nile and eastern equine encephalitis viruses in Southeastern Virginia.

To investigate the occurrence of West Nile virus (WNV) and Eastern equine encephalitis virus (EEE) in southeastern Virginia, the Bureau of Laboratories at the Norfolk Department of Public Health (NDPH) analyzed mosquito pools and the sera of sentinel chickens from the southeastern Virginia area each year from 2000 to 2004. Mosquito pool supernatants were screened for the presence of viral RNA by conventional reverse transcription polymerase chain reaction (RT-PCR) and Taqman RT-PCR with the i-Cycler. Mosquito pools were also tested for virus activity by Vero cell culture. The primary enzootic vector of WNV was Culex (Cx.) pipiens and that of EEE was Culiseta (Cs.) melanura. During the five-year surveillance period, the peak minimum infection rates (MIRs) of WNV and EEE in these mosquito species were 2.7 (2002) and 0.9 (2001), respectively. In 2003, the MIRs in Cs. melanura for WNV and EEE were 0.24 and 0.56, respectively; and the MIR for WNV in Cx. pipiens was 0.64. In 2004, Cs. melanura was less active in the WNV transmission cycle (MIR = 0.07) than was Cx. pipiens (MIR = 1.8), and Cs. melanura was the only vector for EEE (MIR = 0.37). The trend was for EEE activity to peak in July; WNV activity peaked in August. Sentinel-chicken sera were tested for IgM antibodies, and peak IgM seroconversions to these arboviruses were recorded in August 2003 for WNV and in July 2003 for EEE. In 2004, the highest IgM seroconversions to EEE occurred later in August. The overall trend of arbovirus activity was greater in 2003 than in 2004.

Varicella-zoster virus open reading frame 21, which is expressed during latency, is essential for virus replication but dispensable for establishment of latency.

Varicella-zoster virus (VZV) open reading frame 21 (ORF21) is one of at least five VZV genes expressed in latently infected human and rodent ganglia. To determine whether ORF21 is required for latent and lytic infection, we deleted 99% of ORF21 from the viral genome. The ORF21 deletion mutant virus could be propagated only in a cell line expressing the ORF21 protein. Insertion of the herpes simplex virus type 1 (HSV-1) homolog of VZV ORF21, HSV-1 UL37, into the ORF21 deletion mutant failed to complement the mutant for growth in cell culture. Inoculation of cotton rats with the ORF21 deletion virus resulted in latent infection in numbers of animals similar to those infected after inoculation with the parental virus. The mean numbers of latent VZV genomes were similar in animals infected with parental and ORF21 deletion viruses. Transcription of ORF63, another latency-associated gene, was detected in ganglia from similar numbers of animals infected with the mutant and parental viruses. Thus, ORF21 is the first VZV gene expressed during latency that has been shown to be dispensable for the establishment of latent infection.