Mixed infections with respiratory viruses have become frequent at poultry farm business in Egypt. The pandemic of highly pathogenic avian influenza virus (HPAIV) H5N1 (clade 2.2.1.2) has been spreading through the country since 2006. In 2016, the Spread of HPAIV H5N8 (clade 2.3.4.4b) to Egypt via wild birds into commercial poultry flocks made overburden on the economic situation of poultry business. The surveillance efforts conducted by the governmental veterinary authority GOVS showed dominating infection of H5N8 with rare spread of H5N1 in poultry market sector. Thus, the current study try to figure out whether a replication competence exhibited between the two highly pathogenic avian influenza viruses HPAIV H5N1 and H5N8 through designation an in-vitro induced infection of H5N8 and H5N1 in  specific pathogen free embryonated chicken eggs (SPF- ECE).
A concentration of 10³ EID50  of HPAIV H5N1 and H5N8 were either sequentially or simultaneously inoculated into the allantoic cavity of SPF-ECE at 48 h of incubation, followed by the collection of allantoic fluid. A quantitative reverse transcription real-time polymerase chain reaction was used to determine the quantitative replication of viral RNA copies of neuraminidase gene of both AIV strains.
A replication competence was observed according to which viral strain firstly inoculated. Whereas, H5N1 RNA titers was reduced by ≥4log₁₀ when H5N8 firstly inoculated. Vice versa H5N8 RNA titers were reduced by ≥4log₁₀ when H5N1 firstly inoculated. The interference impact of H5N1 was more powerful than that of H5N8 when simultaneous inoculation of both viruses was applied.
In conclusion, quantitative evaluation of mixed inoculation of AIV strains exhibited replication competence of the first virus inoculation on the account of the succeeding one regardless of viral strain. However, H5N8 showed lower replication intensity according to the neuraminidase gene evaluation when both H5N1 and H5N8 were simultaneously inoculated in SPF ECE.