الفهرس 
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Abstract
6.The present study concerned with a trial for isolation of LSD virus from skin lesions of cattle on CAM of ECE as well as molecular characterization of the virus isolate using PCR and sequencing with establishment of a phylogenetic tree between the local LSDV strain and the previously recovered Egyptian isolates.
LSD suspected cattle showed multiple localized or generalized skin nodules. Skin lumps varied in size but were mostly 1-3 cm in diameter and were quite hard in consistency and affected the full thickness anywhere on the animal body.
Skin nodules from clinically diseased cattle were collected as aseptically as possible and used for isolation of the virus. Field samples were inoculated in CAMs. The CAMs were hemorrhagic with congestion in blood vessels appeared by the 1st passage then become more pronounced after the 2nd, 3rd and 4th passages.
The isolated virus was identified on histopathological basis by examination of infected CAMs. characteristic eosinophilic intracytoplasmic inclusion bodies characteristic for poxviridae appeared. These inclusions were suggestive for the presence of LSDV as a member of capripoxviruses replicating in the cytoplasm.
For molecular characterization, an extracted viral DNA from pooled CAMs of ECE was used for detection of LSDV with PCR. The extracted viral DNA from pooled CAMs of ECE was amplified using PCR.Primers specific for GPCR gene amplified a 554 bp product from LSDV genome confirming positivity of the sample for presence of LSD viral genome while primers set targeting RP030 gene amplified a 172 bp product from LSDV genome confirming positivity of the sample for presence of LSD viral genome. Tissue culture adapted SPPV vaccinal strain amplified a (152 bp) which was easily distinguishable relative to the LSDV amplicons.
The local LSDV strain was sequenced and submitted to GenBank with their designation as; (LSDV/ Egy-BSU/2018) under accession number of (MK552139).
The phylogenetic tree grouped viruses of LSD, sheep pox and goat pox viruses separately, even though the obtained strain resources were related to each other and to the other LSDVs.
In the current study GPCR gene nucleotide and their deduced amino acid sequences of LSDV were analyzed by comparing with respective reference sequences obtained from the GenBank.
Nucleotide and amino acids identities were found to be 99.6, 99.4 respectively with LSDV/Egypt/Sharqia/2014 (MF156211.1) and LSDV/Egypt/Beni-suef-1/2014 (KJ561442.1) due to presence of one silent mutation (A111T) and one non-silent mutation (G86A) that led to amino acid substitution (S29N) (Serine into Asparagine), while nucleotide and amino acids identities were found to be 99.2, 98.7 respectively with LSDV/Mansoura/Egypt/2011 (KP071936.1) due to presence of two silent mutations (A156G) and (T246C) and two non-silent mutations (A82T) and (T247C) that led to amino acid substitution (I28F) (Isoleucine into Phenylalanine) and (C83R) (Cysteine into Arginine) respectively.
To study the relationship between theCaPVs, a representative phylogenetic analysis using the neighbor-joining method was carried out on nucleic acid sequences.
Results of phylogenetic and multi-sequence analyses revealed a high degree of similarity between LSDV isolates from different locations with minimal genetic variation. Therefore, from these obtained results we recommend the use of the original Neethling LSDV strain of cattle as an essential strain for controlling of LSD instead of Kenyan sheep pox or goat pox strains.