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RNAi targeting of West Nile virus in mosquito midguts promotes virus diversification.

Brackney DE, Beane JE, Ebel GD - PLoS Pathog. (2009)

Bottom Line: Our data demonstrate that WNV infection in orally exposed vector mosquitoes induces the RNAi pathway and that regions of the WNV genome that are more intensely targeted by RNAi are more likely to contain point mutations compared to weakly targeted regions.These results suggest that, under natural conditions, positive selection of WNV within mosquitoes is stronger in regions highly targeted by the host RNAi response.Further, they provide a mechanistic basis for the relative importance of mosquitoes in driving WNV diversification.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of New Mexico School of Medicine, Albuquerque, NM, USA.

ABSTRACT
West Nile virus (WNV) exists in nature as a genetically diverse population of competing genomes. This high genetic diversity and concomitant adaptive plasticity has facilitated the rapid adaptation of WNV to North American transmission cycles and contributed to its explosive spread throughout the New World. WNV is maintained in nature in a transmission cycle between mosquitoes and birds, with intrahost genetic diversity highest in mosquitoes. The mechanistic basis for this increase in genetic diversity in mosquitoes is poorly understood. To determine whether the high mutational diversity of WNV in mosquitoes is driven by RNA interference (RNAi), we characterized the RNAi response to WNV in the midguts of orally exposed Culex pipiens quinquefasciatus using high-throughput, massively parallel sequencing and estimated viral genetic diversity. Our data demonstrate that WNV infection in orally exposed vector mosquitoes induces the RNAi pathway and that regions of the WNV genome that are more intensely targeted by RNAi are more likely to contain point mutations compared to weakly targeted regions. These results suggest that, under natural conditions, positive selection of WNV within mosquitoes is stronger in regions highly targeted by the host RNAi response. Further, they provide a mechanistic basis for the relative importance of mosquitoes in driving WNV diversification.

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Size and Abundance of Small RNA Reads Mapping to the WNV Genome.The total abundance of sRNAs mapping to the WNV genome based on sRNA length. Black and grey bars correspond to 7 and 14 days post infection, respectively.
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ppat-1000502-g001: Size and Abundance of Small RNA Reads Mapping to the WNV Genome.The total abundance of sRNAs mapping to the WNV genome based on sRNA length. Black and grey bars correspond to 7 and 14 days post infection, respectively.

Mentions: To define the population of small RNAs in mosquito midguts, small RNA (sRNA) libraries from WNV-infected and uninfected midguts after a 7 or 14 day extrinsic incubation period (EIP) were subjected to high-throughput sequencing. The total number of sRNA reads obtained from each of the four libraries sequenced ranged from 3.7 to 4.9 million (Table 1). To determine the presence of viRNAs, sRNA reads were aligned to the WNV genome. There were 2,544 (1,701 unique) and 4,419 (2,629 unique) reads that aligned to the WNV genome following a 7 or 14 day EIP, respectively, where unique reads represent individual viRNAs that code for a specific nucleotide sequence. As expected, these reads had a mean and mode length of 21 nt (Figure 1, Table 1). In addition, the average quality scores [Illumina scores, scaled from 1 (minimum) to 40 (maximum)] were extremely high (90% of the reads >36) and greater than 80% of these reads perfectly aligned with the viral genome (Table 1). A small population of sRNAs from the uninfected mosquito midguts aligned to the WNV genome. A high proportion of these reads required 2 mismatches in order to align to the WNV genome; 50% and 86% at 7 and 14 days post bloodmeal, respectively, (Table 1).


RNAi targeting of West Nile virus in mosquito midguts promotes virus diversification.

Brackney DE, Beane JE, Ebel GD - PLoS Pathog. (2009)

Size and Abundance of Small RNA Reads Mapping to the WNV Genome.The total abundance of sRNAs mapping to the WNV genome based on sRNA length. Black and grey bars correspond to 7 and 14 days post infection, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2698148&req=5

ppat-1000502-g001: Size and Abundance of Small RNA Reads Mapping to the WNV Genome.The total abundance of sRNAs mapping to the WNV genome based on sRNA length. Black and grey bars correspond to 7 and 14 days post infection, respectively.
Mentions: To define the population of small RNAs in mosquito midguts, small RNA (sRNA) libraries from WNV-infected and uninfected midguts after a 7 or 14 day extrinsic incubation period (EIP) were subjected to high-throughput sequencing. The total number of sRNA reads obtained from each of the four libraries sequenced ranged from 3.7 to 4.9 million (Table 1). To determine the presence of viRNAs, sRNA reads were aligned to the WNV genome. There were 2,544 (1,701 unique) and 4,419 (2,629 unique) reads that aligned to the WNV genome following a 7 or 14 day EIP, respectively, where unique reads represent individual viRNAs that code for a specific nucleotide sequence. As expected, these reads had a mean and mode length of 21 nt (Figure 1, Table 1). In addition, the average quality scores [Illumina scores, scaled from 1 (minimum) to 40 (maximum)] were extremely high (90% of the reads >36) and greater than 80% of these reads perfectly aligned with the viral genome (Table 1). A small population of sRNAs from the uninfected mosquito midguts aligned to the WNV genome. A high proportion of these reads required 2 mismatches in order to align to the WNV genome; 50% and 86% at 7 and 14 days post bloodmeal, respectively, (Table 1).

Bottom Line: Our data demonstrate that WNV infection in orally exposed vector mosquitoes induces the RNAi pathway and that regions of the WNV genome that are more intensely targeted by RNAi are more likely to contain point mutations compared to weakly targeted regions.These results suggest that, under natural conditions, positive selection of WNV within mosquitoes is stronger in regions highly targeted by the host RNAi response.Further, they provide a mechanistic basis for the relative importance of mosquitoes in driving WNV diversification.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, University of New Mexico School of Medicine, Albuquerque, NM, USA.

ABSTRACT
West Nile virus (WNV) exists in nature as a genetically diverse population of competing genomes. This high genetic diversity and concomitant adaptive plasticity has facilitated the rapid adaptation of WNV to North American transmission cycles and contributed to its explosive spread throughout the New World. WNV is maintained in nature in a transmission cycle between mosquitoes and birds, with intrahost genetic diversity highest in mosquitoes. The mechanistic basis for this increase in genetic diversity in mosquitoes is poorly understood. To determine whether the high mutational diversity of WNV in mosquitoes is driven by RNA interference (RNAi), we characterized the RNAi response to WNV in the midguts of orally exposed Culex pipiens quinquefasciatus using high-throughput, massively parallel sequencing and estimated viral genetic diversity. Our data demonstrate that WNV infection in orally exposed vector mosquitoes induces the RNAi pathway and that regions of the WNV genome that are more intensely targeted by RNAi are more likely to contain point mutations compared to weakly targeted regions. These results suggest that, under natural conditions, positive selection of WNV within mosquitoes is stronger in regions highly targeted by the host RNAi response. Further, they provide a mechanistic basis for the relative importance of mosquitoes in driving WNV diversification.

Show MeSH
Related in: MedlinePlus