Avian Flu, Wigtownshire: EIR release

Information requested

On your website it states "Highly pathogenic avian influenza (HPAI) H5N1 was confirmed at a premises near Stranraer, Wigtownshire, Dumfries and Galloway on 28 January 2023." (https://www.gov.scot/publications/avian-influenza-outbreaks/).

Under the Freedom of Information (Scotland) Act 2002, I require the following information:

1. Please specify which diagnostic test was used for this purpose (eg, PCR/antigen, etc).
2. How many tests were carried out?
3. How many were positive vs how many were negative?
4. Please provide test particulars (eg, if PCR was used, please specify the precise test (make + model, etc) along with manufacturer's data sheet, and also the cycle threshold of the test(s); OR any other datas relevant to the test(s) in question.
5. Which laboratory (or laboratories) was (or were) used for these tests?
6. Which confirmatory tests were employed on top of the above tests to confirm diagnosis?

As the information you have requested is ‘environmental information’ for the purposes of the Environmental Information (Scotland) Regulations 2004 (EIRs), we are required to deal with your request under those Regulations. We are applying the exemption at section 39(2) of the Freedom of Information (Scotland) Act 2002 (FOISA), so that we do not also have to deal with your request under FOISA.

This exemption is subject to the ‘public interest test’. Therefore, taking account of all the circumstances of this case, we have considered if the public interest in disclosing the information outweighs the public interest in applying the exemption. We have found that, on balance, the public interest lies in favour of upholding the exemption, because there is no public interest in dealing with the same request under two different regimes. This is essentially a technical point and has no material effect on the outcome of your request.

Response

The answer to your question is:

1. Please specify which diagnostic test was used for this purpose (eg, PCR/antigen, etc).
   Real-time PCR and virus isolation in embryonated fowls’ eggs (EFEs) was used.
2. How many tests were carried out?
   A suite of four real-time PCR tests comprising (i) generic influenza A virus screening PCR; (ii) specific PCR for detection of high pathogenicity H5 (‘HP H5’) avian influenza virus; (iii) specific PCR for detection of avian influenza viruses of subtype N1 and (iv) generic Newcastle disease virus screening PCR were each used to test the 45 original clinical samples submitted from 22 birds. The 45 clinical samples comprised an oropharyngeal and a cloacal swab taken from each bird (i.e. 22 sets of paired swabs) and pooled brain tissue taken from the carcasses of two of the birds. Virus isolation in EFEs was also carried out on the pooled brain tissue.
   
3. How many were positive vs how many were negative?
   All 22 birds were positive for HPAIV H5N1 as at least one swab from each bird (plus the brain sample collected from two of the birds) tested positive by all three PCR tests (i), (ii) and (iii). In total, of the 135 (3 x 45) PCR tests run for detection of influenza A virus (i), HP H5 (ii) and N1 (iii), respectively, 126 test results were positive while only nine were negative. All 45 samples were negative by the generic Newcastle disease virus screening PCR.
4. Please provide test particulars (eg, if PCR was used, please specify the precise test (make + model, etc) along with manufacturer's data sheet, and also the cycle threshold of the test(s); OR any other datas relevant to the test(s) in question.
   For the PCR testing, AriaMx (Agilent) real-time instruments were used following automated extraction of nucleic acid from each test sample using a KingFisher Flex (ThermoFisher) robot.
   
   Each of the four PCR tests is run for forty cycles through to completion. A positive sample for the generic influenza A virus screening PCR, specific PCR for detection of high pathogenicity H5 (‘HP H5’) avian influenza virus and the specific PCR for detection of avian influenza viruses of subtype N1 is noted by a fluorescent signal appearing on or before thirty-six cycles of amplification. Any signal appearing after thirty-six cycles of amplification or no signal appearing at all will signify a negative test result for that sample in either of these PCR assays. A positive sample for the generic Newcastle disease virus screening PCR is noted by a fluorescent signal appearing on or before thirty-seven cycles of amplification. Therefore, with this PCR, any signal appearing after thirty-seven cycles of amplification or no signal appearing at all will signify a negative test result.
   
   The main publications describing the PCR tests used for generic influenza A virus screening PCR, specific PCR for detection of high pathogenicity H5 (‘HP H5’) avian influenza virus and the specific PCR for detection of avian influenza viruses of subtype N1 are as follows:
   
   Nagy A, Černíková L, Kunteová K, Dirbáková Z, Thomas SS, Slomka MJ, Dán Á, Varga T, Máté M, Jiřincová H, Brown IH. A universal RT-qPCR assay for "One Health" detection of influenza A viruses. PLoS One. 2021 Jan 20;16(1):e0244669. doi: 10.1371/journal.pone.0244669. PMID: 33471840; PMCID: PMC7817021. James J, Seekings AH, Skinner P, Purchase K, Mahmood S, Brown IH, Hansen RDE, Banyard AC, Reid SM. Rapid and sensitive detection of high pathogenicity Eurasian clade 2.3.4.4b avian influenza viruses in wild birds and poultry. J Virol Methods. 2022 Mar;301:114454. doi: 10.1016/j.jviromet.2022.114454. Epub 2022 Jan 6. PMID: 34998830.
   
   Payungporn S, Chutinimitkul S, Chaisingh A, Damrongwantanapokin S, Buranathai C, Amonsin A, Theamboonlers A, Poovorawan Y. Single step multiplex real-time RT-PCR for H5N1 influenza A virus detection. J Virol Methods. 2006 Feb;131(2):143-7. doi: 10.1016/j.jviromet.2005.08.004. Epub 2005 Sep 23. PMID: 16183140.
   
   The Newcastle disease virus screening PCR details can be found in the following publication:
   
   Sutton DA, Allen DP, Fuller CM, Mayers J, Mollett BC, Londt BZ, Reid SM, Mansfield KL, Brown IH. Development of an avian avulavirus 1 (AAvV-1) L-gene real-time RT-PCR assay using minor groove binding probes for application as a routine diagnostic tool. J Virol Methods. 2019 Mar;265:9-14. doi: 10.1016/j.jviromet.2018.12.001. Epub 2018 Dec 21. PMID: 30579921.
5. Which laboratory (or laboratories) was (or were) used for these tests?
   The tests were performed in the Virology Department at APHA-Weybridge.
   
6. Which confirmatory tests were employed on top of the above tests to confirm diagnosis?
   Whole-genome sequencing was also used to confirm the above diagnosis and the results generated from all laboratory tests are scrutinised in close conjunction with the clinical presentation seen on the IP while it is under restriction.

About FOI

The Scottish Government is committed to publishing all information released in response to Freedom of Information requests. View all FOI responses at http://www.gov.scot/foi-responses.