DNA sequencing and phylogenetic analysis
Currently, Calf guard® vaccine genotype (G6P1) had accession number (MW714568) and 5 different field genotypes (2 G6P5, 1 G10P5, 1G10P? and 1G10P11) that had accession numbers (MW656245, MW751820, MW751824, MW751825 and MW751826) were subjected to DNA sequencing. We observed that at nucleotide level in field genotypes, two G10P5 and G10P? sequences were 100% identical with each other, two G6P5 sequences were 100% identical with each other and there was no significant similarity between sequence of G10P11 with sequences of G6P5, G10P5 and G10P? (Table 1). NCBI BLAST search revealed that G10P5 and G10P? sequences with published sequences in GenBank database showed overall identities of 94.91- 99.66% in nucleotide level, 2 G6P5 sequences with published sequences in GenBank database showed overall identities of 94.01- 98.10% in nucleotide level and G10P11 sequence with published sequences in GenBank database showed overall identities of 94.76- 98.75% in nucleotide level. The phylogenetic analysis of G10P5 and G10P? isolates showed close cluster with G10 isolates of Sharkia governorate, Egypt (Fig. 1). The phylogenetic analysis of 2 G6P5 isolates showed close cluster with VP4 gene of Rotavirus isolate of Dakahlia governorate, Egypt (accession number MK961093) (Fig. 2). The phylogenetic analysis G10P11 isolate showed close cluster with VP4 gene of Rotavirus isolate of Dakahlia governorate, Egypt (accession number MK961092) (Fig. 3).
Molecular comparison between detected and typed Rotaviruses' genotypes with other genotypes of common commercial vaccines
In our result, NCBI BLAST search revealed that G10P5 and G10P? sequences with G10P11 Scour guard vaccine strain showed overall identities of 97.13% in nucleotide level. NCBI BLAST search revealed that G10P5 and G10P? sequences with G6P1 Scour guard vaccine strain and G6P5 Rotavac vaccine strain showed overall identities of 77.62% and 77.31% in nucleotide level, respectively. NCBI BLAST search revealed that G10P11 sequence with G10P11 Scour guard vaccine strain showed overall identities of 95.45% in nucleotide level. NCBI BLAST search revealed that two G6P5 sequences with G6P5 Rotavac vaccine strain showed overall identities of 89.06% in nucleotide level. NCBI BLAST search revealed that there was no significant similarity between different field strains sequences in the present study and sequence of Calf guard® vaccine strain (G6P1) and sequence of VP4 gene of G6P1 of Scour guard vaccine at nucleotide level. NCBI BLAST search revealed that there was no significant similarity between sequence of different field strains (G6P5 and G10P11) in the current study with sequence of VP7 genes of G6P1 and G10P11 of Scour guard vaccine and G6P5 of Rotavac vaccine at nucleotide level. NCBI BLAST search revealed that there was no significant similarity between sequences of different field strains (G6P5, G10P5 and G10P?) in the present study with sequence of VP4 gene of G10P11 of Scour guard vaccine at nucleotide level. NCBI BLAST search revealed that there was no significant similarity between sequences of different field strains (G10P5, G10P? and G10P11) in the current study with sequence of VP4 gene of G6P5 of Rotavac vaccine at nucleotide level (Table 2). Nucleotide sequences in the partial VP7 and VP4 genes of BRVA field strains were compared with those of BRVA vaccine strains. Dissimilarity of nucleotide sequences was observed in several positions in vaccines strains in comparison to field strains (Fig. 4, 5 and 6).
Table (1): Comparison between detected and typed Rotaviruses' genotypes by nucleotide identity percentage
Field strains
|
One G6P5 (VP4 gene)
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One G10P5 (VP7 gene)
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One G10P? (VP7 gene)
|
|
nucleotide identity (%)
|
|
nucleotide identity (%)
|
One G6P5 (VP4 gene)
|
100%
|
*
|
*
|
One G10P5 (VP7 gene)
|
*
|
100%
|
100%
|
One G10P11(VP4 gene)
|
*
|
*
|
*
|