Filogenetska i molekularna analiza gena hemaglutinina virusa ptičje gripe h9n2 izoliranih u jatima peradi u Maroku između 2016. i 2018. godine

Avian influenza viruses of the H9N2 subtype continue to spread in wild birds and poultry worldwide. Infection with H9N2 avian influenza virus was detected for the first time in Morocco in January 2016. In this study, a total of 105 organ and tracheal swab samples from 21 broiler farms in Morocco were collected from July 2016 to October 2018 for H9N2 screening. The suspicion of disease was based on severe respiratory signs such as sneezing, coughing, rales and gasping, while H9N2 virus infection was confirmed by real-time RT-PCR. Hemagglutinin (HA) genes of four isolates were amplified by conventional RT-PCR, sequenced, and aligned for phylogenetic analyses. Among the 21 flocks, 48% (10/21) were qRT-PCR positive for H9, with the cycle threshold values ranging from18.6 to 34.8. The maximum similarity in nucleotide and protein sequences (96-98%) was observed between the Moroccan viruses and an H9 virus isolated from broiler chickens in 2017 in Burkina Faso (A/chicken/BurkinaFaso/17RS93-19/2017) and from a layer chicken in the United Arab Emirates in 2015 (A/chicken/Dubai/D2506/2015). The HA genes revealed the close relationship between the four Moroccan viruses, with 97.9%-99.9% nucleotide identity. Phylogenetic analysis showed that the Moroccan viruses belonged to the G1 lineage, and likely originated from the Middle East, as previously reported in 2016.Virusi ptičje gripe H9N2 nastavljaju se širiti u peradi i divljih ptica širom svijeta. Infekcija niskopatogenim virusom influence H9N2 prvi je put otkrivena u Maroku u siječnju 2016. godine. U ovom je istraživanju za probir na H9N2 prikupljeno ukupno 105 organa i obrisaka iz dušnika s 21 farme brojlera od srpnja 2016. do listopada 2018. iz različitih regija Maroka. Sumnja na bolest temeljila se na teškim respiracijskim znakovima kao što su kihanje, kašljanje, hropanje i hripanje, a infekcija virusom H9N2 potvrđena je PCR-om obrnute transkripcije u stvarnom vremenu. Sekvencije gena za hemaglutinin (HA) od četiri izolata amplificirane su pomoću RT-PCR qRT-PCR poravnane za filogenetsku i analizu sličnosti aminokiselina. Od 21 uzorka jata 48 % (10/21) bilo je pozitivno na H9 s pragom broja ciklusa u rasponu od 18,6 do 34,8. Maksimalna sličnost u nukleotidnim i proteinskim sekvencijama (96 -98 %) uočena je između marokanskih virusa i virusa H9 izoliranih iz brojlerskih pilića u 2017. u Burkini Faso (A/piletina/BurkinaFaso/17RS93-19) i od kokošjeg pileta u Ujedinjenim Arapskim Emiratima u 2015. (A/piletina/ Dubai/D2506/2015). HA geni otkrili su blisku vezu između četiriju virusa, s 97,9 % -99,9 % nukleotidnog identiteta. Filogenetska analiza pokazala je da marokanski virusi pripadaju lozi G1 i vjerojatno potječu s Bliskog istoka, kao što je objavljeno 2016. godine

Spatiotemporal Dynamics, Evolutionary History and Zoonotic Potential of Moroccan H9N2 Avian Influenza Viruses from 2016 to 2021

The H9N2 virus continues to spread in wild birds and poultry worldwide. At the beginning of 2016, the H9N2 Avian influenza virus (AIV) was detected in Morocco for the first time; despite the implementation of vaccination strategies to control the disease, the virus has become endemic in poultry in the country. The present study was carried out to investigate the origins, zoonotic potential, as well as the impact of vaccination on the molecular evolution of Moroccan H9N2 viruses. Twenty-eight (28) H9N2 viruses collected from 2016 to 2021 in Moroccan poultry flocks were isolated and their whole genomes sequenced. Phylogenetic and evolutionary analyses showed that Moroccan H9N2 viruses belong to the G1-like lineage and are closely related to viruses isolated in Africa and the Middle East. A high similarity among all the 2016–2017 hemagglutinin sequences was observed, while the viruses identified in 2018–2019 and 2020–2021 were separated from their 2016–2017 ancestors by long branches. Mutations in the HA protein associated with antigenic drift and increased zoonotic potential were also found. The Bayesian phylogeographic analyses revealed the Middle East as being the region where the Moroccan H9N2 virus may have originated, before spreading to the other African countries. Our study is the first comprehensive analysis of the evolutionary history of the H9N2 viruses in the country, highlighting their zoonotic potential and pointing out the importance of implementing effective monitoring systems