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Occurrence of genetic modifications in core, 5'UTR and NS5b of HCV associated with viral response to treatment.

Kanwal S, Mahmood T - Virol. J. (2014)

Bottom Line: At nucleotide and amino acid level, the genetic distance and mutation, number of predicted N-phosphorylation and N-glycosylation sites was higher in group 2 as compared to group 1.It can be concluded that heterogeneity both at nucleotide and amino acid level contributed in developing drug resistant phenotype.Furthermore, prediction of phosphorylation and glycosylation sites could help in targeting the proper sites for drug designing.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Sciences, Faculty of Biological Sciences, Quaid-i-Azam University, Islamabad 45320, Pakistan. tmahmood.qau@gmail.com.

ABSTRACT

Background: It is becoming progressively more understandable that genetic variability of viruses is a major challenge in translating the laboratory findings to clinic. Genetic variability is the underlying cause of variant viral proteins which are not targetable by host immunological machinery.

Methods: 500 patients were enrolled in study and amongst them, 451 patients were followed and categorized into two groups on the basis of their treatment response. Group 1 consisting of the 376 patients exhibited SVR while group 2 comprised 75 patients who were non-responders on the basis of viral load as evidenced by Real-Time PCR. Comparative sequence analysis was done between 75 non-responders and 75 responders (randomly picked from 376) by targeting three genomic regions, 5'UTR, core and NS5B and amplified products were directly sequenced and obtained sequences were cleaned, aligned and submitted to GenBank. Maximum Parsimony (MP) method was used for phylogenetic analysis and dendrograms were dragged using MEGA 5. Heterogeneity at nucleotide and amino acid level was determined using software BioEdit and DNAman while phosphorylation and N-linked glycosylation sites were determined using NetPhos 2.0 and SignalP-NN.

Results: Genotype 3 was prevalent in group 1 whereas non-responders indicated rare genotypes of Pakistan i.e. 4 and 5, genotype 6q and 6v were reported first time from Pakistan in this study. At nucleotide and amino acid level, the genetic distance and mutation, number of predicted N-phosphorylation and N-glycosylation sites was higher in group 2 as compared to group 1. Difference in percentage composition of individual amino acids was noted to be different between the two groups.

Conclusions: It can be concluded that heterogeneity both at nucleotide and amino acid level contributed in developing drug resistant phenotype. Moreover, occurrence of rare genotypes might hurdle the way to positive response of conventional treatment. Furthermore, prediction of phosphorylation and glycosylation sites could help in targeting the proper sites for drug designing.

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Related in: MedlinePlus

Parsimonious phylogenetic tree constructed using core sequences of patients from group both groups with reference sequences from database.
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Fig6: Parsimonious phylogenetic tree constructed using core sequences of patients from group both groups with reference sequences from database.

Mentions: Phylogenetic tree constructed maximum parsimony method (MP) using sequences of 5׳ UTR from both treatment groups exhibiting the mix pattern of evolution of both the treatment groups (Figure 5). Overall the tree was divided into 16 clusters. Cluster 1 and 2 constitute some sequences of group 1 dipicting the early evolution of the sequences from patients who responded positively to therapy. Moreover sequences from group 1 were present in all the clusters of the tree. Majority of the sequences from the group 2 were present in the recently evolved clusters (clusters 12, 13, 14, 15, 16, 19 and 20). Most of the sequences from group 2 make their separate clusters while somewhere in the tree they possess the position along with the reference sequence from the database (clusters 6, 7, 8, 9, 10, 12). The unresolved genotypes were found in close neighbor to genotype 2, 3, 4 and 6 (clusters5, 6 and 17).Evolutionary tree based on core sequences exhibited the similar branching pattern of group 1 and 2. Overall the tree was divided into 15 clusters. Some sequences from group 1 and 2 were found at the base of the tree and exhibited the early evolutionary period (cluster 1). In cluster 2 sequences from group 1 showed close homology with Asian isolates. At some positions throughout the tree topology some of the sequences from both groups were clustered with each other (clusters 4, 5, 10, 12 and 13) while in others clusters they were found in close proximity with reference strains throughout the world (clusters 3, 6, 7, 8, 9, 11, 14 and 15). Three out of 5 unresolved genotypes are present in cluster 13 and in neighbor of genotype 5 while one unresolved strain was found in cluster 15 along with the genotype 1and another unresolved strain took the place in cluster 7 along with genotype 4 (Figure 6).Dendrogram constructed using sequences from NS5B exhibited diverge branching pattern by almost all the sequences from both groups 1 and 2. Tree was divided into 10 clusters. Sequences from group 1 and 2 along with the reference sequences from Europe were positioned at the base of the tree exhibiting the early evolutionary behavior. Most of the sequences from group 2 were depicting the recent evolution (cluster 6, 8, 9 and 10). Some sequences from group 1 and 2 presenting the homology with sequences from Africa, America (North and South) and Europe (cluster 2, 3, 5, 7 and 8). Moreover the unresolved sequences showed homology with genotypes 1, 3, 4 and 5 and 6 (cluster 6, 8, 9 and 10) (Figure 7).Figure 5


Occurrence of genetic modifications in core, 5'UTR and NS5b of HCV associated with viral response to treatment.

Kanwal S, Mahmood T - Virol. J. (2014)

Parsimonious phylogenetic tree constructed using core sequences of patients from group both groups with reference sequences from database.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4289283&req=5

Fig6: Parsimonious phylogenetic tree constructed using core sequences of patients from group both groups with reference sequences from database.
Mentions: Phylogenetic tree constructed maximum parsimony method (MP) using sequences of 5׳ UTR from both treatment groups exhibiting the mix pattern of evolution of both the treatment groups (Figure 5). Overall the tree was divided into 16 clusters. Cluster 1 and 2 constitute some sequences of group 1 dipicting the early evolution of the sequences from patients who responded positively to therapy. Moreover sequences from group 1 were present in all the clusters of the tree. Majority of the sequences from the group 2 were present in the recently evolved clusters (clusters 12, 13, 14, 15, 16, 19 and 20). Most of the sequences from group 2 make their separate clusters while somewhere in the tree they possess the position along with the reference sequence from the database (clusters 6, 7, 8, 9, 10, 12). The unresolved genotypes were found in close neighbor to genotype 2, 3, 4 and 6 (clusters5, 6 and 17).Evolutionary tree based on core sequences exhibited the similar branching pattern of group 1 and 2. Overall the tree was divided into 15 clusters. Some sequences from group 1 and 2 were found at the base of the tree and exhibited the early evolutionary period (cluster 1). In cluster 2 sequences from group 1 showed close homology with Asian isolates. At some positions throughout the tree topology some of the sequences from both groups were clustered with each other (clusters 4, 5, 10, 12 and 13) while in others clusters they were found in close proximity with reference strains throughout the world (clusters 3, 6, 7, 8, 9, 11, 14 and 15). Three out of 5 unresolved genotypes are present in cluster 13 and in neighbor of genotype 5 while one unresolved strain was found in cluster 15 along with the genotype 1and another unresolved strain took the place in cluster 7 along with genotype 4 (Figure 6).Dendrogram constructed using sequences from NS5B exhibited diverge branching pattern by almost all the sequences from both groups 1 and 2. Tree was divided into 10 clusters. Sequences from group 1 and 2 along with the reference sequences from Europe were positioned at the base of the tree exhibiting the early evolutionary behavior. Most of the sequences from group 2 were depicting the recent evolution (cluster 6, 8, 9 and 10). Some sequences from group 1 and 2 presenting the homology with sequences from Africa, America (North and South) and Europe (cluster 2, 3, 5, 7 and 8). Moreover the unresolved sequences showed homology with genotypes 1, 3, 4 and 5 and 6 (cluster 6, 8, 9 and 10) (Figure 7).Figure 5

Bottom Line: At nucleotide and amino acid level, the genetic distance and mutation, number of predicted N-phosphorylation and N-glycosylation sites was higher in group 2 as compared to group 1.It can be concluded that heterogeneity both at nucleotide and amino acid level contributed in developing drug resistant phenotype.Furthermore, prediction of phosphorylation and glycosylation sites could help in targeting the proper sites for drug designing.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Sciences, Faculty of Biological Sciences, Quaid-i-Azam University, Islamabad 45320, Pakistan. tmahmood.qau@gmail.com.

ABSTRACT

Background: It is becoming progressively more understandable that genetic variability of viruses is a major challenge in translating the laboratory findings to clinic. Genetic variability is the underlying cause of variant viral proteins which are not targetable by host immunological machinery.

Methods: 500 patients were enrolled in study and amongst them, 451 patients were followed and categorized into two groups on the basis of their treatment response. Group 1 consisting of the 376 patients exhibited SVR while group 2 comprised 75 patients who were non-responders on the basis of viral load as evidenced by Real-Time PCR. Comparative sequence analysis was done between 75 non-responders and 75 responders (randomly picked from 376) by targeting three genomic regions, 5'UTR, core and NS5B and amplified products were directly sequenced and obtained sequences were cleaned, aligned and submitted to GenBank. Maximum Parsimony (MP) method was used for phylogenetic analysis and dendrograms were dragged using MEGA 5. Heterogeneity at nucleotide and amino acid level was determined using software BioEdit and DNAman while phosphorylation and N-linked glycosylation sites were determined using NetPhos 2.0 and SignalP-NN.

Results: Genotype 3 was prevalent in group 1 whereas non-responders indicated rare genotypes of Pakistan i.e. 4 and 5, genotype 6q and 6v were reported first time from Pakistan in this study. At nucleotide and amino acid level, the genetic distance and mutation, number of predicted N-phosphorylation and N-glycosylation sites was higher in group 2 as compared to group 1. Difference in percentage composition of individual amino acids was noted to be different between the two groups.

Conclusions: It can be concluded that heterogeneity both at nucleotide and amino acid level contributed in developing drug resistant phenotype. Moreover, occurrence of rare genotypes might hurdle the way to positive response of conventional treatment. Furthermore, prediction of phosphorylation and glycosylation sites could help in targeting the proper sites for drug designing.

Show MeSH
Related in: MedlinePlus