One hundred twenty years of koala retrovirus evolution determined from museum skins

Abstract

Although endogenous retroviruses are common across vertebrate genomes, the koala retrovirus (KoRV) is the only retrovirus known to be currently invading the germ line of its host. KoRV is believed to have first infected koalas in northern Australia less than two centuries ago. We examined KoRV in 28 koala museum skins collected in the late 19th and 20th centuries and deep sequenced the complete proviral envelope region from five northern Australian specimens. Strikingly, KoRV env sequences were conserved among koalas collected over the span of a century, and two functional motifs that affect viral infectivity were fixed across the museum koala specimens. We detected only 20 env polymorphisms among the koalas, likely representing derived mutations subject to purifying selection. Among northern Australian koalas, KoRV was already ubiquitous by the late 19th century, suggesting that KoRV evolved and spread among koala populations more slowly than previously believed. Given that museum and modern koalas share nearly identical KoRV sequences, it is likely that koala populations, for more than a century, have experienced increased susceptibility to diseases caused by viral pathogenesis.

Fig. 1.

Variation in the env region of KoRV in five koala museum specimens collected across a century. Nucleotide positions are listed for polymorphisms (generally two character states) detected in each KoRV. Both character states listed were present at each polymorphism listed for each individual. The modern sequence (GenBank DQ174772) matched KoRV sequences from all koala museum specimens, including the first of the listed character states at each of the polymorphic sites shown. The only exception was a fixed nucleotide difference at position 1940, indicated by the symbol “§” in which the modern reference sequence had guanine but all museum sequences had adenine. Nucleotide positions are numbered from the start codon of env. A “d” denotes polymorphisms in which the reference sequence did not match the character state present in GALV. Nonsynonmous nucleotide variation is italicized, with codon number and single letter amino acid codes next to the corresponding nucleotide polymorphism, with the amino acid matching the KoRV reference listed first. An asterisk (*) indicates amino acid changes that could be structurally modeled.

Fig. 2.

Museum KoRV ENV protein structural variation. (A) Structural superimpositions between KoRV and GALV (top panel), or KoRV and PERV (lower panel), show in cartoon representations the overall similarity of the structures, with major differences located at the receptor-binding sites (yellow boxes). These differences (arrows) include the presence or absence of α helices, different angles between the loops, and the presence of additional loops. (B) Distribution of the electrostatic potential on solvent-accessible surfaces of KoRV (top panel), GALV (middle panel), and PERV (lower panel) structures, showing that the major differences in protein charge are located at the receptor-binding sites, indicated by yellow boxes. Additional differences are indicated by arrows. Positively charged groups are blue, negative charges are red, and white gray areas are neutral. (C) The structural model of KoRV ENV proteins showing the positions of four nonsynonymous mutations (upper panel; mutations are also listed in fig. 1). Lower panels show structural alignments at sites with amino acid variants (arrows), focusing on mutations and resultant structural changes. The G189S mutation was predicted to cause a major structural alteration that changed the angle and conformation of a conserved loop of unknown function. W90C and T245A both exchanged a surface amino acid with a partially buried one. Both G187D and G189S were predicted to alter the electrostatic profile of the protein surface (not shown).