Microarray analysis of evolution of RNA viruses: Evidence of circulation of virulent highly divergent vaccine-derived polioviruses

Abstract

Two approaches based on hybridization of viral probes with oligonucleotide microarrays were developed for rapid analysis of genetic variations during microevolution of RNA viruses. Microarray analysis of viral recombination and microarray for resequencing and heterogeneity analysis were able to generate instant genetic maps of vaccine-derived polioviruses (VDPVs) and reveal the degree of their evolutionary divergence. Unlike conventional methods based on cDNA sequencing and restriction fragment length polymorphism, the microarray approaches are better suited for analysis of heterogeneous populations and mixtures of different strains. The microarray hybridization profile is very sensitive to the cumulative presence of small quantities of different mutations, including those that cannot be revealed by sequencing, making this approach useful for characterization of profiles of nucleotide sequence diversity in viral populations. By using these methods, we identified a type-3 VDPV isolated from a healthy person and missed by conventional methods of screening. The mutational profile of the polio strain was consistent with >1 yr of circulation in human population and was highly virulent in transgenic mice, confirming the ability of VDPV to persist in communities despite high levels of immunity. The proposed methods for fine genotyping of heterogeneous viral populations can also have utility for a variety of other applications in studies of genetic changes in viruses, bacteria, and genes of higher organisms. molecular evolution ͉ recombination ͉ population heterogeneity ͉ mutation screening ͉ live vaccine

Application of a Microarray-Based Assay for the Study of Genetic Diversity of West Nile Virus

“…Although the microarray hybridization method provides limited information about the position of mutations in the analyzed genomic region when compared to sequencing, it has the significant advantage of allowing the identification of "hot spots" where random mutations occur within short (a few nucleotides) areas. It also may allow for the detection of those mutations even when they occur at relatively low levels (up to 1%) as in the case of mixtures of quasispecies that cannot be detected by traditional direct sequencing methods (Cherkasova et al, 2003;Leberre at al., 2007). The efficiency of microarrays for identification and discrimination of closely related bacteria and viruses has been previously demonstrated Hsia et al, 2007;Laassri et al, 2003;Nordström et al, 2005;Volokhov et al, 2002;Wade et al, 2004).…”

“…Microarray technology has been used to study gene expression in clinical and biological samples, detect and genotype pathogens (Honma et al, 2007;Wade et al, 2004), detect single base pair mismatches (Anthony et al, 2003;Hacia et al, 1999), design genomic maps (Roerig et al, 2005), and study viral evolution (Cherkasova et al, 2003).…”

Microarray Techniques for Evaluation of Genetic Stability of Live Viral Vaccines

“…Thus, sensitive high-throughput microarray techniques including microarray for resequencing and sequence heterogeneity (MARSH), microarray analysis of viral recombination (MAVR) assay, and microarray for quantitation of known virulent mutations (MQNVM) have been developed and applied as valuable tools to evaluate the genetic stability of live viral vaccines (Cherkasova et al, 2003;Laassri et al, 2011;Laassri et al, 2005;Laassri et al, 2007).…”

“…The MARSH assay was first used to analyze mutations that accumulate in the region coding for VP1, the most variable capsid protein of poliovirus (Cherkasova et al, 2003;Laassri et al, 2005). Later, this microarray approach was expanded to discriminate between vaccinia strains and to evaluate genetic stability of the vaccinia virus Ankara (MVA) B5R gene following propagation of a cloned isolate of MVA in Vero and MRC-5 cell lines (Laassri et al, 2007), to analyze the variability of the structural region of West Nile (WN) virus (Grinev et al, 2008), and to evaluate stability of the entire genome of a WN/Dengue 4 chimeric virus under study as a new candidate of WN vaccine (Laassri et al, 2011).…”

Resolution in forensic microbial genotyping

“…Oligonucleotide micro-arrays are an emerging technology for mutational screening of Human DNA as well as bacteria and viruses [83][84][85][86][87]. They are one of the few potentially practical ways of detecting SNSs over significant fractions of the bacterial genome without direct sequencing.…”