NEW YORK (GenomeWeb) – Researchers have reconstructed the sequence of a hepatitis B virus that infected a 16th century mummy, and have found similarities between it and modern strains.
The mummy, which was excavated from an Italian church about 35 years ago, was initially suspected to be infected with smallpox based on a paleopathological analysis, but researchers from McMaster University were unable to recover any variola virus from the mummy. However, as they reported today in PLOS Pathogens, they were able to uncover HBV DNA that displayed the hallmarks of ancient DNA.
Despite appearing centuries old, McMaster's Hendrik Poinar and his colleagues noted that the mummy's HBV was closely related to modern HBV strains belonging to genotype D. If the virus sample is both genuinely old and similar to modern strains, this indicates that HBV has been infecting humans for centuries. Currently, about 350 million people have chronic HBV infections.
"The more we understand about the behavior of past pandemics and outbreaks, the greater our understanding of how modern pathogens might work and spread, and this information will ultimately help in their control," Poinar said in a statement.
Radiocarbon dating indicated that the mummy, a child who died at about two years of age, was about 439 years old. This timing, the researchers noted, was in line with how the child was dressed, embalmed, and buried.
A previous autopsy uncovered a rash on the child's body, and electron microscopy and immunostaining of pustules suggested the child was possibly infected with smallpox. However, Poinar and his colleagues repeated that analysis and were unable to find variola or hepatitis viral particles. But they extracted DNA from five body sites and enriched those samples for HBV, variola, and mitochondrial DNA.
From these samples, the researchers teased out HBV reads, which they mapped to the HBV reference genome and identified as belonging to the subgenotype D3. Subgenotype D3 has a global distribution and is common in the Mediterranean, the researchers noted. They then constructed a consensus HBV genome 3,182 nucleotides in length, though with a gap in a GC-rich region.
The researchers reported being unable to map any reads to the variola genome. This suggested to Poinar and his colleagues that the rash the child had was Gianotti-Crosti syndrome, a rare HBV symptom, instead of a smallpox rash.
The HBV DNA samples harbored a deamination pattern that the researchers said was expected in an ancient sample, rather than a contaminating modern one. Additionally, they noted that the pattern was similar to what they observed in the mitochondrial DNA they analyzed from the mummy, supporting the notion that the samples are old.
Poinar and his colleagues also constructed phylogenetic trees of the wider HBV population and the D genotype. The Italian sample they analyzed here as well as one previously obtained from a 330-year-old Korean mummy both fell among modern viral populations. The authenticity of the Korean HBV sample is uncertain, as a deamination analysis was not reported, the researchers noted.
Additionally, the Italian HBV sample was grouped among modern HBV D3 subgenotype samples.
This finding, Poinar and his colleagues said, presents an apparent paradox as the samples appear old, but fall within the modern diversity of HBV and its subgenotypes. If the findings hold true, the researchers said that they suggest that HBV genotypes diversified before 1500 CE and that any diversity that has accumulated since has been lost through purifying selection or been masked by multiple substitutions.
It would also indicate that HBV has been associated with human populations for quite some time, they added. "These results have important implications for our understanding of the emergence and evolution of this common viral pathogen," the researchers wrote.