Internal Validation of the ASFV MONODOSE dtec-qPCR Kit for African Swine Fever Virus Detection under the UNE-EN ISO/IEC 17025:2005 Criteria

SIMPLE SUMMARY: African swine fever virus is a pathogen capable of spreading among swine populations, which is usually fatal since no vaccines are currently available on the market. Outbreaks caused by this virus are the reason for massive economic losses on pig farms and are a matter of worry for the swine sector worldwide. Effective and reliable detection of the virus is relevant to prevent uncontrolled contagion, especially when no effective preventive measures can be applied. In order to confront the spread of African swine fever, several rapid real-time polymerase chain reaction detection assays were developed by different research groups. In the present study, we focused on the validation of the ASFV MONODOSE dtec-qPCR kit using reference genetic material provided by national and international reference laboratories, and results were compared with those obtained from using internationally accepted reference methods. The data obtained indicated that the kit can be used in veterinary samples. Technical innovations enable a user-friendly and rapid management of samples, reducing the probability of human errors happening and transport costs. ABSTRACT: African swine fever virus is considered an emerging virus that causes African swine fever, a disease characterised by high mortality and elevated transmission rates and that, as it is for most other viral diseases, cannot be treated with specific drugs. Effective and reliable detection of the virus is relevant to prevent uncontrolled contagion among boar populations and to reduce economic losses. Moreover, animal health laboratories are demanding standardisation, optimisation and quality assurance of the available diagnostic assays. In the present study, the ASFV MONODOSE dtec-qPCR kit was validated following the UNE-EN ISO/IEC 17025:2005 guidelines. Analytical validation terms include in silico and in vitro specificity, sensitivity, efficiency and reliability (repeatability/reproducibility). Diagnostic validation of the method was assessed through the analysis of a total of 181 porcine samples originating from six different matrix types doped with African swine fever virus DNA received from the European reference laboratory for African Swine Fever (INIA-CISA, Madrid, Spain): whole blood, blood serum, kidney, heart, liver and tonsil. Results agreed with those obtained from a reference detection method also based on real-time PCR, endorsed by WOAH, but the ASFV MONODOSE dtec-qPCR kit incorporates some technical innovations and improvements which may benefit end-users. This kit, available worldwide with full analytical and diagnostic validation, can recognise all known ASFV genotypes and brings additional benefits to the current qPCR technology.