Comparison and optimization of detection methods for noroviruses in frozen strawberries containing different amounts of RT-PCR inhibitors. (December 2016)

• Record Type: Journal Article
• Authors: Bartsch, Christina; Szabo, Kathrin; Dinh-Thanh, Mai; Schrader, Christina; Trojnar, Eva; Johne, Reimar
• Is Part Of: Food microbiology. Volume 60(2016:Dec.)
• Page Start: 124
• Page End: 130
• Publication Date: 2016-12
• Subjects: Norovirus -- Strawberries -- Detection method -- PCR inhibitors; Food Microbiology -- Periodicals; Aliments -- Microbiologie -- Périodiques; Food -- Microbiology; Periodicals; Food -- Microbiology -- Periodicals; Food contamination -- Periodicals; 664.001579
• DOI: 10.1016/j.fm.2016.07.005
• Languages: English
• ISSNs: 0740-0020

Abstract

Abstract: Frozen berries have been repeatedly identified as vehicles for norovirus (NoV) transmission causing large gastroenteritis outbreaks. However, virus detection in berries is often hampered by the presence of RT-PCR-inhibiting substances. Here, several virus extraction methods for subsequent real-time RT-PCR-based NoV-RNA detection in strawberries were compared and optimized. NoV recovery rates (RRs) between 0.21 ± 0.13% and 10.29 ± 6.03% were found when five different artificially contaminated strawberry batches were analyzed by the ISO/TS15216-2 method indicating the presence of different amounts of RT-PCR inhibitors. A comparison of five different virus extraction methods using artificially contaminated strawberries containing high amounts of RT-PCR inhibitors revealed the best NoV RRs for the ISO/TS15216 method. Further improvement of NoV RRs from 2.83 ± 2.92% to 15.28 ± 9.73% was achieved by the additional use of Sephacryl ® -based columns for RNA purification. Testing of 22 frozen strawberry samples from a batch involved in a gastroenteritis outbreak resulted in 5 vs. 13 NoV GI-positive and in 9 vs. 20 NoV GII-positive samples using the original ISO/TS15216 method vs. the extended protocol, respectively. It can be concluded that the inclusion of an additional RNA purification step can increase NoV detection by the ISO/TS15216-2 method in frozen berries containing high amounts of RT-PCR inhibitors. Highlights: Frozen strawberries can contain different amounts of RT-PCR inhibitors. Efficiency of norovirus extraction protocols from strawberries varies remarkably. The ISO/TS15216 method shows good performance with frozen strawberries. Additional RNA purification with Sephacryl ® -based columns increases detection rate. Use of these columns is suggested for norovirus detection in berry samples.