Sequence and phylogenetic analysis of FMD virus isolated from two outbreaks in Egypt.

Despite intensive control efforts, Foot and mouth disease (FMD) outbreaks continue to occur regularly in Egypt and resulting in dramatic economic losses to the livestock industry. During 2018 and 2022, FMD was clinically suspected among previously vaccinated cattle in Beheira and Kafr El-Sheikh provinces, Egypt. FMDV RNA was detected in 18 (45%) out of 40 epithelial tissue samples using real-time RT-PCR based on a pan-FMDV primers set. The 2018 outbreak isolates (n = 8) included the FMDV serotypes A and SAT2, whereas all isolates (n = 10) from the 2022 outbreak belonged to the FMDV serotype A. Four selected isolates, designated FMDV/SAT2/EGY/Beheira/2018, FMDV/A/EGY/Kafr El-Sheikh/2018, FMDV/A/EGY/Kafr El-Sheikh/2022 and FMDV/A/EGY/Behiera/2022, were characterized on the basis of partial VP1 gene sequence analysis. The FMDV/SAT2/EGY/Beheira/2018 strain was clustered within the Lib-12 lineage of the topotype VII and shared 79.2-98.4% nucleotide identity with other Egyptian SAT2 strains available in Genbank database. On the other hand, the three FMDV serotype A sequences shared 74.4-99.1% nucleotide identity with each other. Also, they were phylogenetically classified within two distinct topotypes. The FMDV/A/Egy/Kafr El-Sheikh/2018 strain was grouped within the Asian topotype, meanwhile the FMDV/A/EGY/Kafr El-Sheikh/2022 and FMDV/A/EGY/Behiera/2022 strains were grouped together within the genotype IV of the African topotype. Interestingly, the deduced amino acid sequences of the four strains displayed numerous variations in comparison to the vaccine strains currently used in Egypt. In addition, most of these variations were present in prominent antigenic positions in the VP1 protein. These findings raise a crucial need to validate the protective potential of the vaccine strains against the newly emerging FMDV field strains and to update the vaccination strategy accordingly.

Co-circulation of GI.1 and GI.2 genotypes of rabbit hemorrhagic disease virus in Egypt.

Recently, Egypt has experienced an increased incidence of rabbit hemorrhagic disease virus (RHDV) infection even among vaccinated rabbits. The present study estimates the emergence of RHDV in vaccinated (n = 10) and unvaccinated (n = 8) domestic rabbitries in Beheira and Kafr El-Sheikh provinces, Egypt, during the period 2018-2020. A total of 8 out of 18 (44.4%) liver extracts were able to agglutinate human type O RBCs with HA titers ranged from 8 to 12 log2, and then subsequently confirmed for the presence of RHDV RNA using a reverse transcriptase-polymerase chain reaction (RT-PCR). The VP60 gene sequences of three selected isolates, designated Beh-1, Beh-9 and kaf-14, were submitted to the GenBank database and the accession numbers MZ782083 to MZ782085 were assigned, respectively. Phylogenetic analysis revealed that the Kaf-14 isolate was placed into the GI.1 genotype, while the Beh-1 and Beh-9 isolates were grouped into the GI.2 genotype. Overall, the three isolates shared 78.6-98.7%.nucleotide identity with previously published Egyptian sequences. In comparison with the GI.1a Giza2006 vaccine strain, the three isolates exhibited divergence ranging from 4.5 to 17.4% at the amino acid level. Approximately 55.5-87.5% of the amino acid substitutions were located in the P2 subdomain of the VP60 capsid protein which contains the main determinants of antigenicity and cellular recognition. In conclusion, our results provide crucial evidence for the co-circulation of RHDV GI.1 and GI.2 genotypes in Egypt and highlight the antigenic diversity among vaccine and field strains. Therefore, new effective vaccines are urgently required to counter the spread of GI.1 and GI.2 genotypes in Egypt.

First report of seroprevalence and genetic characterization of avian orthoreovirus in Egypt.

Recently, the Egyptian broiler industry has experienced an increased incidence of avian reovirus (ARV) infections. However, to date, no studies have been carried out to investigate the epidemiologic status of ARV infections as well as the genetic characteristics of the currently circulating ARV strains. The present study estimates the seroprevalence of ARV infections in Alexandria, El-Behera, Giza, Kafr El-Sheikh, and Gharbia governorates, Egypt, during the period 2017-2018. A total of 150 serum samples from 15 unvaccinated broiler flocks with suspicious ARV infection were screened using a commercial enzyme-linked immunosorbent assay kit. All the tested flocks were found to be positive for ARV-specific antibodies, and the overall seropositivity rate was 80.6%. Meanwhile, 5 (33.3%) flocks were confirmed for the presence of ARV through a reverse transcription-polymerase chain reaction (RT-PCR) assay based on the σA-encoding gene. Phylogenetic analysis based on the nucleotide sequences of the σA-encoding gene revealed that the obtained ARV isolate, designated EGY1, was grouped in the S1113-like cluster of ARV and displayed 100% and 98.7% nucleotide identity with the Chinese MSO1 isolate and the S1133 vaccine strain, respectively. In addition, amino acid alignments with the S1133 vaccine strain revealed that the σA protein of the EGY1 isolate carried the substitutions G81S and A118V. In conclusion, the present study provides the evidence for a ubiquitous distribution of ARV infection in Egypt as well as represents a starting point for genetic characterization of the currently circulating ARV strains.

Protection conferred by a vaccine derived from an inactivated Egyptian variant of infectious bronchitis virus: a challenge experiment.

The current study investigated the protective efficacy of a formalin-inactivated infectious bronchitis virus (IBV) vaccine derived from the field strain KP729422, which exhibits low S1 spike protein sequence homology (77.1-79.8%) with the currently used vaccine strains in Egypt. Two-week-old, specific-pathogen-free chickens were subcutaneously inoculated with a single dose of the vaccine containing 106.7 50% embryo infective dose (EID50) of the inactivated virus. At 6 weeks of age, the chickens were challenged with 104 EID50 of the same virus strain via the oculonasal route. In comparison with the unvaccinated challenged group, the vaccinated chickens had significantly higher IBV-neutralizing antibody titers and exhibited efficient protection against challenge on the basis of tracheal ciliary activity. However, the challenge virus was recovered from the kidneys and tracheas of these chickens at rates of 40% and 60%, respectively. These findings suggest that a single application of the vaccine may provide sufficient clinical and respiratory protection, but may not ensure complete protection against infection by the challenge virus.

Isolation and Molecular Characterization of Novel Infectious Bronchitis Virus Variants from Vaccinated Broiler Flocks in Egypt.

The present study aimed to determine the molecular characteristics of circulating infectious bronchitis virus (IBV) strains in vaccinated broiler flocks in the Giza and Fayoum governorates. Thirty-four isolates were collected, and egg propagation revealed their ability to induce typical IBV lesions after three to five successive passages. Three selected isolates were identified as IBV using a real-time reverse transcriptase-PCR assay targeted the nucleocapsid (N) gene and further characterized by partial spike (S) gene sequence analysis. Phylogenetic analysis revealed their clustering into two variant groups. Group I consisted of one variant (VSVRI_F3), which had 99.1% nucleotide sequence identity to the Q1 reference strain. Group II consisted of variants VSVRI_G4 and VSVRI_G9, which showed 92.8%-94.3% nucleotide identity with the Egyptian variants Eg/12120S/2012, Eg/12197B/2012, and Eg/1265B/2012. Regarding the deduced amino acid sequence, the three variants had 77.1%-85.2% similarity with the vaccine strains currently used in Egypt. These findings highlight the importance of monitoring the prevalence of IBV variants in vaccinated broiler flocks as well as adopting an appropriate vaccination strategy.