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Summary

Abstract

Introduction

Protocol

Representative Results

Discussion

Acknowledgements

Materials

References

Environment

Detection of Viruses from Bioaerosols Using Anion Exchange Resin

Published: August 22nd, 2018

DOI:

10.3791/58111

1High Plains Intermountain Center for Agricultural Health and Safety, Department of Environmental and Radiological Health Sciences, Colorado State University, 2National Wildlife Research Center, Wildlife Services, Animal and Plant Health Inspection Service, United States Department of Agriculture, 3Western Sydney University, 4Leprino Foods, Inc, 5Department of Food Science and Agricultural Chemistry, McGill University, 6Department of Mechanical Engineering, Colorado State University, 7Department of Animal Science, University of Wyoming

An anion exchange resin-based method, adapted to liquid impingement-based bioaerosol sampling of viruses is demonstrated. When coupled with downstream molecular detection, the method allows for facile and sensitive detection of viruses from bioaerosols.

This protocol demonstrates a customized bioaerosol sampling method for viruses. In this system, anion exchange resin is coupled with liquid impingement-based air sampling devices for efficacious concentration of negatively-charged viruses from bioaerosols. Thus, the resin serves as an additional concentration step in the bioaerosol sampling workflow. Nucleic acid extraction of the viral particles is then performed directly from the anion exchange resin, with the resulting sample suitable for molecular analyses. Further, this protocol describes a custom-built bioaerosol chamber capable of generating virus-laden bioaerosols under a variety of environmental conditions and allowing for continuous monitoring of environmental variables such as temperature, humidity, wind speed, and aerosol mass concentration. The main advantage of using this protocol is increased sensitivity of viral detection, as assessed via direct comparison to an unmodified conventional liquid impinger. Other advantages include the potential to concentrate diverse negatively-charged viruses, the low cost of anion exchange resin (~$0.14 per sample), and ease of use. Disadvantages include the inability of this protocol to assess infectivity of resin-adsorbed viral particles, and potentially the need for the optimization of the liquid sampling buffer used within the impinger.

The purpose of this method is to provide a highly sensitive bioaerosol sampling platform to facilitate molecular detection of negatively-charged viruses from bioaerosols. Microorganisms, including viral particles, can survive in bioaerosols for extended periods of time1. Bioaerosols can travel over relatively long distances and maintain viability and infectivity, as evidenced by an outbreak of Legionnaires' disease that originated from industrial cooling towers located at a distance of 6 km from the affected individuals and resulted in 18 fatalities2. Indirect transmission of viruses to humans mediated by bioaerosols....

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1. Setup of the Bioaerosol Chamber (See Figure 2)

  1. Pre-load the liquid impingers with 20 mL of 0.01 M phosphate buffered saline, pH 7.5 (PBS).
    1. Add 0.5 g of anion exchange resin and suspend within the PBS of one of the liquid impingers, with another liquid impinger serving as a control.
  2. Position liquid impingers in parallel inside the bioaerosol chamber using clamp stands with aerosol inlets facing the nebulizer.
    NOTE: See.......

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Figure 1 demonstrates the principle behind charge-based capture of viruses from bioaerosols via inclusion of resin in liquid-based impingers. Figure 2 shows the setup of the custom-built bioaerosol chamber. Figure 3 describes the steps involved in setting up the aerosolization experiment and measures to ensure quality control. Figure 4 shows amplification curves for qRT-.......

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This protocol outlines a method for sensitive viral capture from bioaerosols using modified liquid impingers. The method is optimized for detection and quantification of the viral load in bioaerosols. The specific modification demonstrated here involves the addition of anion exchange resin to liquid contained within a common liquid impinger. This method was developed for its simplicity in downstream sample processing, whereas other sample processing techniques such as centrifugation, filtration, and precipitation-based m.......

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This work was supported by funding from the CDC/NIOSH High Plains Intermountain Center for Agricultural Health and Safety (5U54OH008085) and the Colorado Bioscience Discovery Evaluation Grant Program (14BGF-16).

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Name Company Catalog Number Comments
Escherichia coli bacteriophage MS2 (ATCC 15597-B1) American Type Culture Collection ATCC 15597-B1
FluMist Quadrivalent AstraZeneca Contact manufacturer Viral constitutents of this vaccine are subject to change on an annual basis
CFX96 Touch Real-Time PCR Detection System Bio-Rad 1855195
Primers and probes Integrated DNA Technologies NA
0.2 µM sterile filter NA NA
1 L pyrex bottles or equivalent NA NA
1 mL pipet tips NA NA
1 mL pipettor NA NA
50 mL serological pipet NA NA
PCR tubes NA NA
Pipet-aid or equivalent NA NA
QIAamp Viral RNA Mini Kit Qiagen 52904
QuantiTect Probe RT-PCR Kit Qiagen 204443
Amberlite IRA-900 chloride form Sigma-Aldrich 216585-500G
Phosphate buffered saline Sigma-Aldrich P5368-10PAK
Water (molecular biology grade) Sigma-Aldrich W4502-1L
Eppendorf DNA LoBind Microcentrifuge Tubes ThermoFisher 13-698-791
Falcon 50 mL Conical Centrifuge Tubes  ThermoFisher 14-432-22
Falcon Polypropylene Centrifuge Tubes ThermoFisher 05-538-62
SuperScript III Platinum One-Step qRT-PCR Kit w/ROX ThermoFisher 11745100
SKC Biosampler 20 mL, 3-piece glass set SKC Inc. 225-9593
Vac-u-Go sample pumps SKC Inc. 228-9695
Collison nebulizer (6-jet) BGI Inc. NA
HEPA capsule PALL 12144
Q-TRAK indoor air quality monitor 8554 TSI Inc. NA
Alnor velometer thermal anemometer AVM440-A TSI Inc. NA
SidePak AM510 personal aerosol monitor TSI Inc. NA
Bioaerosol chamber NA NA

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