HBV sequences used in this study.

<p>^† Details of the GenBank Accession Numbers for all HBV sequences are provided in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132533#pone.0132533.s004" target="_blank">S1 Table</a>.</p><p>^‡ Five HBV/C sequences initially used in phylogenetic tree construction representing HBV/C7, C9, C10, C15, and C16 were not used in nucleotide divergence analysis because only single complete genome isolates were available for each of these subgenotypes.</p><p>^§ Core immune epitope analysis used sequences that cover the C gene region. Compared to the sequences used in the surface immune epitope analysis, only 16 HBV/C Asia sequences were used again in the core immune epitope analysis, while all S gene sequences from HBV/C Papua Pacific and the 87 HBV/C Indonesia of this study did not qualify for the core immune epitope analysis.</p><p>HBV sequences used in this study.</p

Phylogenetic tree of HBV/C isolates from different countries in East and Southeast Asia, and Papua-Pacific.

<p>A Bayesian phylogenetic tree analysis based on complete genome sequences showed that isolates from various subgenotypes (C1-C16) are clearly grouped into two major clusters, consistent with their geographical origins. Seven HBV/C subgenotypes (C1, C2, C5, C7, C8, C9, and C10) from East and Southeast Asia, and one (C14) from Papua (<i>light highlight</i>) were well-separated from those six subgenotypes (C6, C11, C12, C13, C15, and C16) from Papua, and from one subgenotype (C3) from Pacific, the more east region of the Papua (<i>dark highlight</i>). Although the root of subgenotype C3 phylogenetically is distanced from the subgenotypes of Papua, the isolate geographic origin, the immune epitope characteristics of surface and core proteins, and the HBsAg subtype gradient distribution showed these HBV/C3 isolates to be close to Papua subgenotypes. Therefore, the Papua and the Pacific subgenotypes are classified together into Papua-Pacific type. The diversification of the Asian type from the Papua-Pacific type started from Papua of Indonesia to the east. The other subgenotype, HBV/C4, was distanced from other subgenotypes. In this analysis, one strain (GQ358157) from Papua reported as C6 in our previous study [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0132533#pone.0132533.ref023" target="_blank">23</a>] grouped into C12. We redefine this strain as a member of HBV/C12.</p

HBcAg amino acid motifs in B and T-cell epitopes of Asia and Papua-Pacific HBV/C isolates.

<p>For the sake of clarity, this figure was not drawn to scale. Among 15 amino acid positions examined within HBcAg immune epitopes of 143 isolates, we identified I/V at position c59 as the essential variation that classified HBV/C subgenotypes into two major clusters, the Asian and the Papua-Pacific (p-value <0.001; data not shown). HBV/C4 and C14 showed similar variation in most amino acids examined, with C4 and C14 having cI59 and cV59, respectively.</p

Distribution of HBsAg subtypes and HBV genotypes/subgenotypes of 271 HBV/C isolates according to their country/geographical origins in East/Southeast Asia and Papua-Pacific.

<p>^{# including 37 published complete genome sequences and 87 newly generated in this study; N. Caledonia: New Caledonia; PNG: Papua New Guinea.}</p><p>Distribution of HBsAg subtypes and HBV genotypes/subgenotypes of 271 HBV/C isolates according to their country/geographical origins in East/Southeast Asia and Papua-Pacific.</p

Distribution of HBV/C subtypes in the East and Southeast Asia and the Papua-Pacific.

<p>This study identified a west-to-east gradient in the distribution of HBsAg subtypes with <i>adrq+</i> (<i>red</i>) prominent in East-Southeast Asia and <i>adrq-</i> (<i>pink</i>) in the Pacific region (Vanuatu, Fiji, Tonga, and Kiribati). Interestingly, together with <i>adrq+</i>, <i>adrq-</i>indeterminate sA159/sA177 and a new pattern of <i>adrq-</i>indeterminate sV159/sV177 identified in this study were found in Papua and PNG, respectively, suggesting that the molecular admixture of HBV/C, particularly for subtype evolution, occurred in Papua and PNG with both <i>adrq-</i>indeterminate forms (<i>yellow</i>) as the transitional patterns.</p

Mean percentage nucleotide divergence of the complete genome between HBV/C subgenotypes.

<p>The total number of HBV/C isolates examined for each subgenotype is shown in bracket. Other existing subgenotypes (C7, C9, C10, C15, and C16) were not included in the genetic distance calculation since only single isolate was available for each subgenotype. Intrasubgenotype divergences are shown in bold.</p

The distinct pathways in which gene ontology clusters 88 of the genes with decreased expression normalizing during treatment.

<p>Significant Panther biological processes were detected with 88 down-regulated genes; type 2 interferon expression is decreased in this group.</p

Differentially expressed genes in common between active TB patients versus controls and TB-infected versus uninfected mice.

<p>Significant Panther biological processes were detected from 85 out of the 121 over-represented genes common between the two models.</p

Ingenuity pathway analysis of the key genes identified.

<p>Ingenuity pathway analysis of the genes differentially regulated and common to the three systems described above (in orange). Network of genes in the interferon 1 signaling pathways that were found to be common in active TB patients, BCG-infected THP-1 cells at 20 hours and mice infected with active TB are illustrated. The lines in between genes represent known interactions, with solid lines representing direct interactions and dashed lines representing indirect interactions.</p

The distinct pathways in which gene ontology clusters 56 of the genes with increased expression normalizing during treatment.

<p>Significant Panther biological processes was detected for 56 up-regulated genes, type 1 interferon expression is increased in this group.</p