Characterization and Genetic Analysis of Recent and Emergent Virulent Newcastle Disease Viruses in Egypt

Abstract

Extensive vaccination against Newcastle disease virus (NDV) induced more selective immune pressure from hosts that enhanced the evolutionary process of NDV. Herein, we characterized 13 recently isolated NDV isolates from vaccinated chicken flocks during 2014–2017. Sequence analysis of F gene showed the presence of 112RRQKRF117 velogenic cleavage motif in 11 isolates, whereas the other two isolates (Ck/ME3/Eg/16 and Ck/ME5/Eg/16) showed the monobasic motif 112GGRQGRL117. Interestingly, isolate Ck/ME5/Eg/16 showed 100% and 99.82% nucleotide identity with the LaSota F gene hypervariable region and full‐length gene, respectively. On the other hand, isolate Ck/ME3/Eg/16 revealed natural recombination with strains NDV/Ck/Egypt/3/2006 and NDV/Teal/VRLCU/Egypt/2015 that indicates re‐emergence of that old strain. Interestingly, all 13 isolates showed high intracerebral pathogenicity (ICPI) and mean death time (MDT) despite the presence of lentogenic motif in both Ck/ME5/Eg/16 and Ck/ME3/Eg/16 isolates. Comparative analysis of F antigenic epitopes in our isolates with other published sequences from Egypt revealed high sequence conservation; recent isolates had one fixed amino acid substitution (K78R) and a novel V168I substitution, whereas a D170N substitution was detected in older strains (NDV‐EG‐35‐2014 and NDV‐KFR‐B7‐2012). Taken together, our results support the first isolation of virulent NDV isolates with a lentogenic motif; isolate Ck/ME5/Eg/16 might be generated in nature from LaSota live vaccine, whereas isolate Ck/ME3/Eg/16 is emergent from NDV/Ck/Egypt/3/2006. We conclude that the current diagnostic evaluation of the virulence of NDV isolates by characteristic amino acid residues at the F protein cleavage site is insufficient. There is a need to link virologic and epidemiologic data together, and routine and emergency LaSota vaccination protocols should be carefully and optimally applied, with regards to the timing and presence of co‐infecting agents in the field.

Genomic and antigenic properties of Newcastle Disease virus genotypes 2.XX1 and 2.VII from Egypt do not point to antigenic drift as a driving force of spread

“…All viruses from the year 2006 (NDV/chicken/Egypt/2-4/2006; FJ969393, FJ969394, FJ969395) belong to genotype 2.II but are of velogenic pathotype with ICPI values between 1,6 to 1.8 and a have corresponding proteolytic polybasic cleavage site (RRQKR*FIG). The re-isolation of vaccine type virus in line with earlier description (Abolnik et al, 2004;Nagy et al, 2020) and it is conceivable, that emergency vaccination in flocks with suspected ND might lead to such double infections. Those circumstances have been considered by the OIE (OIE, 2018) when defining criteria for virulence: The definition includes the sequence information of the proteolytic cleavage site, but has the amendment "Failure to demonstrate the characteristic pattern of amino acid residues as described above would require characterization of the isolated virus by an ICPI test".…”

“…Sanger-sequence of isolate R1973/11 pointed to avirulent pathotype, but the ICPI result clearly revealed a velogenic pathotype. Such apparently contradicting results have been published before (Nagy et al, 2020;Tan et al, 2008), but were not further resolved. After observing such contradicting results, we plaque-cloned both isolates and were able to separate two different NDV: beside genotype 2.XXI virus, a vaccine type virus (genotype 2.II) was obtained.…”

“…In the brain multicycle replication is restricted to virulent NDV due to the cleavability of the F-protein (Nagai et al, 1976;Rott, 1979) and thus brain passage lead to enrichment of virulent NDV. In case of contradicting results as described by others (Nagy et al, 2020;Tan et al, 2008) sequencing of re-isolated virus from the brain is indicated.…”

“…Continuously growing phylogenic diversity points to the evolution of virus populations that display a genetic distance to vaccine type NDV genotype 2.II. This observation has been taken as indication for antigenic vaccine mismatch and a suspected cause for vaccine failure (Dimitrov et al, 2017;Nagy et al, 2020). In this model new NDV strains emerge because of an change in antigenic sites relevant for protection.…”

Efficacy of the Newcastle Disease Virus Genotype VII.1.1-Matched Vaccines in Commercial Broilers

“…Developing nervous manifestations following recovery from respiratory and digestive illness has been always associated with velogenic NDV in non-vaccinated broilers or in cases of improperly vaccinated infected flocks [43,44]. The data that were obtained from this study highlight that birds who were vaccinated with a G-II inactivated vaccine (G-1) showed the highest level of nervous manifestations (8%), while the birds who were vaccinated with the either the genotype VII rNDV or Autogenous NDV (G2 and G3) did not show any nervous system manifestations, suggesting a high level of protection in the G-2 and G-3 birds.…”