PMC7802106_01
Laboratory timeline for an adult presenting with acutely deranged liver function tests in the setting of HBV infection.. Anti-HBc IgM and IgG were both present throughout the timeline. Blood for whole genome sequencing was taken at presentation (week 0). Elastography score improved to 25.3kPa 6 months later then down to 20.7kPa at one year after initial presentation.
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ALT 41 IU/L, BR 40mumol/L ( Figure 1)).
Maximum likelihood phylogenetic tree of HBV sequence derived from an adult male presenting acutely with HBV infection.. Consensus HBV sequence for patient 1745 is shown (blue) alongside representative genotype and subtype reference sequences, and all full-length HBV sequences from Pakistan (green) and Malaysia (red) from GenBank. Reference sequences
: A (X02763), B (GQ205440), C (KP017269), D1 (clade 1): KP322600, D1 (clade 2): JN040807, D2: KR905424, D3: KP322602, D5: KP322603, D4/7: FJ692533 (D4 and D7 cluster closely and this sequence is representative of the clade including both genotypes). Bootstrap replicates of 1000 generated using MEGA7. Bootstrap values of >70 are shown. Scale bar shows substitutions per site. The sequences in GenBank from Pakistan mapped to either subtype D1 (6/7 sequences) or D3 (1/7 sequences) whilst Malaysian sequences map to subtype D2 (3/3 sequences).
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The consensus sequence was 3182 nucleotides long and mapped to genotype D, clustering within subtype D1, making it most closely related to other published sequences from Pakistan ( Figure 2). In this deep sequence dataset, 2.5% (80/3182) of nucleotide positions had a Shannon entropy score of >0.1 ( Figure 2B).
HBV genome map showing sequencing coverage, diversity and deletions for a genotype D sequence derived from a patient with acute biochemical hepatitis.. In each case, the nucleotide position is shown on the x-axis, with the approximate positions of corresponding genes indicated in the bar at the top (adapted from McNaughton
et al.
).
A: Illumina data showing read coverage (log scale) at each nucleotide site along the HBV genome. The drop in coverage around nt 1000-1600 corresponds to the single stranded portion of the genome.
B: Shannon entropy calculated for each site along the genome. Shaded areas represent non-overlapping sections of the genome and the horizontal dotted line isolates the top six most diverse sites.
C: Locations of minority variant deletions, with a threshold indicated at >=10% to identify the deletions most likely to be relevant. Deletions shown at the start of the genome are likely to represent an artefact of the mapping (short reads derived from a circular genome have been mapped onto a linear construct).
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After de-duplication, the median coverage was 18,069 reads per site ( Figure 3A). In this deep sequence dataset, 2.5% (80/3182) of nucleotide positions had a Shannon entropy score of >0.1 ( Figure 3B). Minority variant deletions occurring in >=10% of viral sequencing reads were detected at nt 1419-1426 and nt 1860-1865 in the RT/X-gene and pre-core genes respectively ( Figure 3C).
PMC7802106_02