Food and Environmental Virology

, Volume 8, Issue 1, pp 70–78 | Cite as

Evaluation of a Porcine Gastric Mucin and RNase A Assay for the Discrimination of Infectious and Non-infectious GI.1 and GII.4 Norovirus Following Thermal, Ethanol, or Levulinic Acid Plus Sodium Dodecyl Sulfate Treatments

  • Olamide T. Afolayan
  • Cathy C. Webb
  • Jennifer L. Cannon
Original Paper

Abstract

Human noroviruses (NoVs) are a major source of foodborne illnesses worldwide. Since human NoVs cannot be cultured in vitro, methods that discriminate infectious from non-infectious NoVs are needed. The purpose of this study was to evaluate binding of NoV genotypes GI.1 and GII.4 to histo-blood group antigens expressed in porcine gastric mucin (PGM) as a surrogate for detecting infectious virus following thermal (99 °C/5 min), 70 % ethanol or 0.5 % levulinic acid (LV) plus 0.01 or 0.1 % sodium dodecyl sulfate (SDS) sanitizer treatments and to determine the limit of detection of GI.1 and GII.4 binding to PGM. Treated and control virus samples were applied to 96-well plates coated with 1 µg/ml PGM followed by RNase A (5 ng/µl) treatment for degradation of exposed RNA. Average log genome copies per ml (gc/ml) reductions and relative differences (RD) in quantification cycle (Cq) values after thermal treatment were 1.77/5.62 and 1.71/7.25 (RNase A) and 1.73/5.50 and 1.56/6.58 (no RNase A) for GI.1 and GII.4, respectively. Treatment of NoVs with 70 % EtOH resulted in 0.05/0.16 (GI.1) and 3.54/10.19 (GII.4) log reductions in gc/ml and average RD in Cq value, respectively. LV (0.5 %) combined with 0.1 % SDS provided a greater decrease of GI.1 and GII.4 NoVs with 8.97 and 8.13 average RD in Cq values obtained, respectively than 0.5 % LV/0.01 % SDS. Virus recovery after PGM binding was variable with GII.4 > GI.1. PGM binding is a promising surrogate for identifying infectious and non-infectious NoVs after capsid destruction, however, results vary depending on virus strain and inactivation method.

Keywords

Norovirus Porcine gastric mucin Ethanol Levulinic acid Sodium dodecyl sulfate Thermal inactivation Surrogate 

Supplementary material

12560_2015_9219_MOESM1_ESM.docx (43 kb)
Supplementary material 1 Supplementary Fig. 1 Experimental design for PGM plus RNase A assay for thermal treated and non-treated GI.1 and GII.4 (Sydney) viruses. PGM, CBS, and RNase A added to wells of microtiter plate as listed in rows A-H. Sample/controls added to wells as listed in columns 1-5. Shaded wells were not used in the assay. Three replicate trials were performed. (DOCX 42 kb)
12560_2015_9219_MOESM2_ESM.docx (69 kb)
Supplementary material 2 Supplementary Fig. 2. Experimental design for PGM plus RNase A assay for LV/SDS treated and non-treated GI.1 and GII.4 (New Orleans) viruses. PGM, CBS, and RNase A added to wells of microtiter plate as listed in rows A-H. Sample/controls added to wells as listed in columns 1-12. Shaded wells were not used in the assay. Three replicate trials were performed. (DOCX 68 kb)
12560_2015_9219_MOESM3_ESM.docx (41 kb)
Supplementary material 3 Supplementary Fig. 3. Experimental design for PGM plus RNase A assay for EtOH treated and non-treated GI.1 and GII.4 (Sydney) viruses. PGM, CBS, and RNase A added to wells of microtiter plate as listed in rows A-H. Sample/controls added to wells as listed in columns 1-12. Shaded wells were not used in the assay. Two replicate trials were performed. (DOCX 41 kb)

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Olamide T. Afolayan
    • 1
  • Cathy C. Webb
    • 1
  • Jennifer L. Cannon
    • 1
  1. 1.Department of Food Science and Technology, Center for Food SafetyUniversity of GeorgiaGriffinUSA

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