{"title":"Comparison of Extraction Methods for the Detection of Tick-Borne Encephalitis Virus RNA in Goat Raw Milk and Cream Cheese","authors":"Irene Müller, Nadine Althof, Bernd Hoffmann, Christine Klaus, Katja Schilling-Loeffler, Alexander Falkenhagen, Reimar Johne","doi":"10.1007/s12560-022-09535-y","DOIUrl":null,"url":null,"abstract":"<div><p>Infection with the tick-borne encephalitis virus (TBEV) can cause meningitis, meningoencephalitis and myelitis in humans. TBEV is an enveloped RNA virus of the family <i>Flaviviridae</i>, which is mostly transmitted via tick bites. However, transmission by consumption of virus-contaminated goat raw milk and goat raw milk products has also been described. Only a few methods have been reported for the detection of TBEV in food so far. Here, we compare different virus extraction methods for goat raw milk and goat raw milk cream cheese and subsequent detection of TBEV-RNA by RT-qPCR. Langat virus (LGTV), a naturally attenuated TBEV strain, was used for artificial contamination experiments. Mengovirus and the human coronavirus 229E were compared to assess their suitability to serve as internal process controls. Out of three tested extraction protocols for raw milk, sample centrifugation followed by direct RNA extraction from the aqueous interphase yielded the best results, with a recovery rate (RR) of 31.8 ± 4.9% for LGTV and a detection limit of 6.7 × 10<sup>3</sup> LGTV genome copies/ml. Out of two methods for cream cheese, treatment of the samples with TRI Reagent® and chloroform prior to RNA extraction showed the best RR of 4.7 ± 1.6% for LGTV and a detection limit of 9.4 × 10<sup>4</sup> LGTV genome copies/g. RRs of Mengovirus and LGTV were similar for both methods; therefore, Mengovirus is suggested as internal process control virus. The developed methods may be useful for screening or surveillance studies, as well as in outbreak investigations.</p></div>","PeriodicalId":563,"journal":{"name":"Food and Environmental Virology","volume":"15 1","pages":"32 - 42"},"PeriodicalIF":4.1000,"publicationDate":"2022-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s12560-022-09535-y.pdf","citationCount":"2","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Food and Environmental Virology","FirstCategoryId":"97","ListUrlMain":"https://link.springer.com/article/10.1007/s12560-022-09535-y","RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"ENVIRONMENTAL SCIENCES","Score":null,"Total":0}
引用次数: 2
Abstract
Infection with the tick-borne encephalitis virus (TBEV) can cause meningitis, meningoencephalitis and myelitis in humans. TBEV is an enveloped RNA virus of the family Flaviviridae, which is mostly transmitted via tick bites. However, transmission by consumption of virus-contaminated goat raw milk and goat raw milk products has also been described. Only a few methods have been reported for the detection of TBEV in food so far. Here, we compare different virus extraction methods for goat raw milk and goat raw milk cream cheese and subsequent detection of TBEV-RNA by RT-qPCR. Langat virus (LGTV), a naturally attenuated TBEV strain, was used for artificial contamination experiments. Mengovirus and the human coronavirus 229E were compared to assess their suitability to serve as internal process controls. Out of three tested extraction protocols for raw milk, sample centrifugation followed by direct RNA extraction from the aqueous interphase yielded the best results, with a recovery rate (RR) of 31.8 ± 4.9% for LGTV and a detection limit of 6.7 × 103 LGTV genome copies/ml. Out of two methods for cream cheese, treatment of the samples with TRI Reagent® and chloroform prior to RNA extraction showed the best RR of 4.7 ± 1.6% for LGTV and a detection limit of 9.4 × 104 LGTV genome copies/g. RRs of Mengovirus and LGTV were similar for both methods; therefore, Mengovirus is suggested as internal process control virus. The developed methods may be useful for screening or surveillance studies, as well as in outbreak investigations.
期刊介绍:
Food and Environmental Virology publishes original articles, notes and review articles on any aspect relating to the transmission of pathogenic viruses via the environment (water, air, soil etc.) and foods. This includes epidemiological studies, identification of novel or emerging pathogens, methods of analysis or characterisation, studies on survival and elimination, and development of procedural controls for industrial processes, e.g. HACCP plans. The journal will cover all aspects of this important area, and encompass studies on any human, animal, and plant pathogenic virus which is capable of transmission via the environment or food.