Tracking echovirus eleven outbreaks in Guangdong, China: a metatranscriptomic, phylogenetic, and epidemiological study

Figures

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  Figure 1.. Metatranscriptomics analysis on different samples types from the three fatal cases. (A) The workflow for metatranscriptomics NGS sequencing and sequencing data analysis. Anal (B) and oral swabs (C) were sampled from Cases 1 and 3 in autopsy diagnosis. The transcriptomes abundance of different microbe families in Cases 1 and 3 was investigated by CCMetagen. The relative coverage and depth at each position of E11 genome is shown by aligning sequencing data from Cases 1 and 3 to the E11 reference genome (KY981558); (D) Serial blood samples were collected from the fatal Case 5 at different time points (marked with black dots). The dynamic change of transcriptomes abundance of different microbe families was shown in left panel. The relative coverage and depth mapping to E11 genome were shown from Day 8. Micrococci and Picornavirus which were commonly identified in all types of samples with a relative high abundance were marked with asterisk.

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  Figure 2.. Environmental surveillance and seroprevalence of E11. (A) The number of E11 isolates detected each month by cell-culture isolation from 2009 to 2012 and June 2018 to May 2019. To clearly illustrate the seasonal pattern and the increase in E11 circulation between 2009 and 2012, the number of E11 isolates from 2009 to 2012 were marked with black dots of a gradient color and presented between May to next June for each season. The mean number of E11 isolates from 2009 to 2012 and number of E11 isolates from 2018 to 2019 were marked with green line and magenta line, respectively. (B) Seroprevalence of neutralizing antibodies to E11 in different age groups of population in Guangdong, China. The number of serums tested for each group was shown in brackets.

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  Figure 3.. Phylogenetic analysis of E11 VP1 genes. (A) ML tree of VP1 genes was constructed including the sixty-six new E11 VP1 gene sequences identified in this study (marked in orange bars on the left). To provide a background of the diversity of E11 lineages in China, the E11 sequences collected from other regions of China (1994–2016) were highlighted with a green bar. Black circles indicate bootstrap support >0.9 at the root node of selected clade. The Guangdong subclade of E11 including the outbreaks strains and retrospectively detected strains between 2017 and 2019 were enlarged in the box (B) twenty-seven clinical cases from five different hospitals (H1–H4) were highlighted with different color dots according to their administration hospitals. The E11 strains sampled from sewage each month from January 2018 to May 2019 are marked with grey dots. (C) Median-joining haplotype network constructed with clinical sequences from H1 to H4 and the ARI outbreak with the same color scheme in (B).

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  Figure 4.. Recombination analysis of E11 genomes. (A) Bootscanning recombination analysis of E11 strains and other closely related enterovirus B genotypes. The outbreak representative strain (Accession No. MN597939) was used as query sequence. The dashed line indicates >70% bootstrap support. The genome structure of the E11 was annotated according to the E11 reference strain KY981558. ML trees were construct separately for the 5’UTR-3A, 2B-3B (B) and 5’UTR-3A, 3C-3’UTR (C) parts of the E11 genome and genomes from other closely related enterovirus genotypes. The E11 genomes generated in this study (n = 32) and collected from public database (n = 41) were highlighted with black circles and light grey dots, respectively. The different parts of the genome from the same virus were connected with a line. In the phylogeny of 2B-3B and 3C-3’UTR, other enterovirus genotypes clustered with E11 suggesting potential recombination, and name of these enterovirus genotypes were noted on the right.