Research Design and Methods The main objective of this study is to identify and determine airborne emission and subsequent surface burden patterns of RSV in naturally infected patients.
Patient Information:
Patients seen at the Emergency Department( ED) or admitted to the hospital with respiratory symptoms and diagnosed with RSV are eligible for enrollment. The results of routine in-house laboratory diagnostic tests such as the Respiratory Viral Panel (RVP) will be used to identify patients.
Setting:
Wake forest baptist medical center (WFBMC) is an 885 bed tertiary care teaching hospital. There are approximately 36,000 inpatient admissions and \>89,000 ED visits annually (pediatric/adult). Air sampling is conducted in patient rooms under turbulent airflows (6 total air changes/hour) at approximately 20°C temperature and 40% Relative Humidity. Air is filtered by ANSI/ASHRAE 52.2 compliant endfilters with a minimum efficiency reporting value of 15.
Sampling:
One nasopharyngeal swab will be obtained from each subject. The sample will be used to confirm RSV infection and determine the amount of the pathogen carried by the individual. Samples will be inoculated into Becton Dickinson Universal Viral Transport System (VTM, BD Diagnostics, Franklin Lakes, NJ) for real-time reverse transcriptase PCR (rRT-PCR) analysis.
One-time air samples will be collected using six-stage Andersen air sampling devices and settle plates at three locations, head of bed/chair two feet away from patient head, foot of bed/chair eight feet away from head, and at the exit of the patient room (11):
Samples will be collected in viral transport media (VTM), supplemented with RNA stabilization buffer (Qiagen AVL buffer) and carrier RNA and then stored at -80C. Carrier RNA will serve as both a carrier and an internal control by containing genomic RNA of the plant tobacco mosaic virus. A 540 ul volume of RNA sample will be purified with the QIAmp Viral RNA Mini Extraction Kit and concentrated approximately 10-fold. RSV serotypes A and B as well as the control viral tobacco mosaic virus RNA will be detected by reverse transcription and quantitative PCR. Additionally, nNasopharyngeal swab with also be used for cultures with inoculation in petri dish containing RSV specific media.
The surface burden of RSV will be assessed by placement of open standard Petri dishes filled with VTM at the three air sampling locations. The covers of the Petri dishes will be removed at the beginning of and closed after the air sampling session. Selected environmental surfaces will be sampled with sterile swabs moistened with VTM or TSB broth. A two square inch area will be swabbed once at three high-touch locations daily: head of the bed hand rail, middle of food tray table, and a table/horizontal surface at back of room (approximately 10 feet away past foot of bed). The swabs will be vortexed for 30 seconds, and processed as described for RSV detection in air samples.
During the sampling health care professional (HCP) and study staff will wear respirators (N-95) to reduce the risk of external contamination due to potential RSV carrier status. The respirators will be collected and a two square inch midsection of the respirator will be cut out using sterile scissors. The cut-outs are placed in 2mLs VTM or TSB broth and frozen. After thawing, samples are vortexed/agitated, and further processed as described for the RSV samples.
