Identification and Characterization of Sindbis Virus RNA-Host Protein Interactions.

Arthropod-borne viruses, such as the members of the genus Alphavirus, are a significant concern to global public health. As obligate intracellular pathogens, RNA viruses must interact with the host cell machinery to establish and complete their life cycles. Despite considerable efforts to define the host-pathogen interactions essential for alphaviral replication, an unbiased and inclusive assessment of alphaviral RNA-protein interactions has not been undertaken. Moreover, the biological and molecular importance of these interactions, in the full context of their molecular function as RNA-binding proteins, has not been fully realized. The data presented here introduce a robust viral RNA-protein discovery method to elucidate the Sindbis virus (SINV) RNA-protein host interface. Cross-link-assisted mRNP purification (CLAMP) assessment revealed an extensive array of host-pathogen interactions centered on the viral RNAs (vRNAs). After prioritization of the host proteins associated with the vRNAs, we identified the site of protein-vRNA interaction by a UV cross-linking and immunoprecipitation sequencing (CLIP-seq) approach and assessed the consequences of the RNA-protein binding event of hnRNP K, hnRNP I, and hnRNP M in regard to viral infection. Here, we demonstrate that mutation of the prioritized hnRNP-vRNA interaction sites effectively disrupts hnRNP-vRNA interaction. Correlating with disrupted hnRNP-vRNA binding, SINV growth kinetics were reduced relative to wild-type parental viral infections in vertebrate and invertebrate tissue culture models of infection. The molecular mechanism leading to reduced viral growth kinetics was found to be dysregulated structural-gene expression. Collectively, this study further defines the scope and importance of the alphavirus host-pathogen vRNA-protein interactions.IMPORTANCE Members of the genus Alphavirus are widely recognized for their potential to cause severe disease. Despite this recognition, there are no antiviral therapeutics, or safe and effective vaccines, currently available to treat alphaviral infection. Alphaviruses utilize the host cell machinery to efficiently establish and complete their life cycle. However, the extent and importance of host-pathogen RNA-protein interactions are woefully undercharacterized. The efforts detailed in this study fill this critical gap, and the significance of this research is 3-fold. First, the data presented here fundamentally expand the scope and understanding of alphavirus host-pathogen interactions. Second, this study identifies the sites of interaction for several prioritized interactions and defines the contribution of the RNA-protein interaction at the molecular level. Finally, these studies build a strategy by which the importance of the given host-pathogen interactions may be assessed in the future, using a mouse model of infection.

Making a difference: APIC chapters introducing public health graduate students to the infection prevention profession.

Health care workforce recruitment into the field of infection prevention and control (IPC) is imperative to address attrition and maintain safe patient care. Association for Professionals in Infection Control (APIC) Badger Chapter in Wisconsin strategically aligned with their state's academic master of public health program to introduce students to the IP profession and offered scholarships to Wisconsin's annual infection prevention and control conference. Four scholarships were awarded from 2021 to 2022. Student feedback of the experience was positive and resulted in 2 participants becoming employed IPs.

Optimized infection control practices augment the robust protective effect of vaccination for ESRD patients during a hemodialysis facility SARS-CoV-2 outbreak.

BACKGROUND: While dialysis patients are at greater risk of serious SARS-CoV-2 complications, stringent infection prevention measures can help mitigate infection and transmission risks within dialysis facilities. We describe an outbreak of 14 cases diagnosed in a hospital-based outpatient ESRD facility over 13 days in the second quarter of 2021, and our coordinated use of epidemiology, viral genome sequencing, and infection control practices to quickly end the transmission cycle. METHODS: Symptomatic patients and staff members were diagnosed by RT-PCR. Facility-wide screening utilized SARS-CoV-2 antigen tests. SARS-CoV-2 genome sequences were obtained from residual diagnostic specimens. RESULTS: Of the 106 patients receiving dialysis in the facility, 10 were diagnosed with SARS-CoV-2 infection, as was 1 patient support person. Of 3 positive staff members, 2 were unvaccinated and had provided care for 6 and 4 of the affected patients, respectively. Sequencing demonstrated that all cases in the cluster shared an identical B.1.1.7./Alpha substrain. Attack rates were greatest among unvaccinated patients and staff. Vaccine effectiveness was 88% among patients. CONCLUSIONS: Prompt recognition of an infection cluster and rapid intervention efforts successfully ended the outbreak. Alongside consistent adherence to core infection prevention measures, vaccination was highly effective in reducing disease incidence and morbidity in this vulnerable population.

Outbreak or pseudo-outbreak? Integrating SARS-CoV-2 sequencing to validate infection control practices in a dialysis facility.

BACKGROUND: The COVID-19 pandemic poses a particularly high risk for End Stage Renal Disease (ESRD) patients so rapid identification of case clusters in ESRD facilities is essential. Nevertheless, with high community prevalence, a series of ESRD patients may test positive contemporaneously for reasons unrelated to their shared ESRD facility. Here we describe a series of 5 cases detected within 11 days in November 2020 in a hospital-based 32-station ESRD facility in Southwest Wisconsin, the subsequent facility-wide testing, and the use of genetic sequence analysis to evaluate links between cases. METHODS: Four patient cases and one staff case were identified in symptomatic individuals by RT-PCR. Facility-wide screening was conducted using rapid SARS-CoV-2 antigen tests. SARS-CoV-2 genome sequences were obtained from residual diagnostic specimens. RESULTS: Facility-wide screening of 47 staff and 107 patients identified no additional cases. Residual specimens from 4 of 5 cases were available for genetic sequencing. Clear genetic differences proved that these contemporaneous cases were not linked. CONCLUSIONS: With high community prevalence, epidemiological data alone is insufficient to deem a case cluster an outbreak. Cluster evaluation with genomic data, when available with a short turn-around time, can play an important role in infection prevention and control response programs.

A Prospective, Randomized Comparison of Duodenoscope Reprocessing Surveillance Methods.

Duodenoscope use in healthcare facilities has been associated with transmission of multidrug resistant pathogens between patients. To assist healthcare facilities in monitoring the quality of their duodenoscope reprocessing procedures and limit patient risk of infection, the Centers for Disease Control and Prevention (CDC) deployed voluntary interim duodenoscope sampling and culturing surveillance protocols in 2015. Though the interim methods were widely adopted, alternative surveillance protocols were developed and implemented at individual institutions. Here, we compared two sampling methods-the 2015 CDC interim protocol and an alternative protocol developed by the University of Wisconsin Hospitals and Clinics (UWHC). We hypothesized that the UWHC protocol would detect a higher incidence of bacterial contamination from reprocessed duodenoscopes. A total of 248 sampling events were performed at UWHC. The CDC protocol (n = 129 sampling events) required culturing samples collected from each duodenoscope after brushing its terminal end and flushing its lumen with sterile water. The UWHC protocol (n = 119 sampling events) required culturing samples collected from each duodenoscope after swabbing its elevator, immersing its terminal end into broth and flushing its lumen with saline. With the CDC method, 8.53% (n = 11) of the duodenoscopes sampled were positive for bacterial growth with 15 isolates recovered. Using the UWHC method, 15.13% (n = 18) of cultures were positive for bacterial growth with 20 isolates recovered. The relative risk of identifying a contaminated duodenoscope using the CDC interim method, however, was not different than when using the UWHC protocol. Mean processing time (27.35 and 5.11 minutes, p < 0.001) and total cost per sample event ($17.87 and $15.04) were lower using the UWHC method. As the UWHC protocol provides similar detection rates as the CDC protocol, the UWHC method is useful, provided the shorter processing time and lower cost to perform.

Occurrence of Amblyomma americanum (Acari: Ixodidae) and Human Infection With Ehrlichia chaffeensis in Wisconsin, 2008-2015.

Because of the increasing incidence of human ehrlichiosis in Wisconsin, we assessed reports of human infections by Ehrlichia chaffeensis and the distribution of its vector, the lone star tick (Amblyomma americanum (L.)). From 2008 through 2015, 158 probable and confirmed human cases of E. chaffeensis infections were reported to the Wisconsin Department of Health Services. Five cases without travel history outside of Wisconsin were confirmed as E. chaffeensis by polymerase chain reaction. Surveillance for the vector occurred from 2008 through 2015 and was based on active and passive methods, including examination of white-tailed deer, collections from live-trapped small mammals, submissions of ticks removed from wild and domestic animals through the Wisconsin Surveillance of Animals for Ticks (SWAT) program, digital or physical submissions by the public to the University of Wisconsin Insect Diagnostic or Medical Entomology laboratories, and active tick dragging. More than 50 lone star ticks (46 adults, 6 nymphs, and 1 larva) were identified. Lone star ticks were more commonly found in south central Wisconsin, particularly in Dane County, where discovery of more than one life stage in a single year indicates possible establishment.