Emergence of porcine delta-coronavirus pathogenic infections among children in Haiti through independent zoonoses and convergent evolution

Abstract

Coronaviruses have caused three major epidemics since 2003, including the ongoing SARS-CoV-2 pandemic. In each case, coronavirus emergence in our species has been associated with zoonotic transmissions from animal reservoirs ^1,2 , underscoring how prone such pathogens are to spill over and adapt to new species. Among the four recognized genera of the family Coronaviridae - Alphacoronavirus, Betacoronavirus, Deltacoronavirus, Gammacoronavirus , - human infections reported to date have been limited to alpha- and betacoronaviruses ³ . We identify, for the first time, porcine deltacoronavirus (PDCoV) strains in plasma samples of three Haitian children with acute undifferentiated febrile illness. Genomic and evolutionary analyses reveal that human infections were the result of at least two independent zoonoses of distinct viral lineages that acquired the same mutational signature in the nsp15 and the spike glycoprotein genes by convergent evolution. In particular, structural analysis predicts that one of the changes in the Spike S1 subunit, which contains the receptor-binding domain, may affect protein's flexibility and binding to the host cell receptor. Our findings not only underscore the ability of deltacoronaviruses to adapt and potentially lead to human-to-human transmission, but also raise questions about the role of such transmissions in development of pre-existing immunity to other coronaviruses, such as SARS-CoV-2.

Figure 1:

Non-specific CPEs formed by plasma from patient 0081–4 in Vero E6 cells. a) Mock-infected Vero E6 cells, 11 days port-inoculation with phosphate-buffered saline. b) Vero E6 cells 11 days post-inoculation with plasma from patient 0081–4. Original magnification at 200X

Figure 2 |. Maximum likelihood tree of PDCoV strains.

The tree was inferred from 109 PDCoV full genome sequences, including four sparrow CoV genomes for outgroup rooting (accession numbers MG812375, MG812376, MG812377, and MG812378, from Chen et al., 2018) using the best fitting nucleotide substitution model (see Methods). For display purposes, outgroup sequences were removed from the figure. Vertical branches are scaled in number of nucleotide substitutions per site according to the bar on the left of the tree. Bootstrap values > 75% (1000 replicates) are indicated along supported branches. The table at the bottom of the tree shows nucleotide % dissimilarity (and total number of nucleotide differences in parenthesis) between Haitian strains and their closest non-Haitian relatives, which are labelled in tree by progressive letters according to the legend on the right.

Figure 3 |. Bayesian maximum clade credibility (MCC) tree of PDCoV strains.

The MCC tree was inferred from a subset of 94 full genome strains that displayed sufficient temporal signal for molecular clock calibration. Branch lengths were scalded in time, according to the bar at the bottom, by enforcing a strict molecular clock and using sampling dates to estimate PDCoV evolutionary rate. Circles at internal nodes indicate high posterior probability (PP) support >0.9. The table on the left shows the inferred time of the most recent common ancestor (TMRCA) between Haitian strains and their closest phylogenetic relative, with 95% high posterior density intervals (95%HPD).

Figure 4 |. Analysis of conserved amino acids in Hu-PDCoV strains.

a) Amino acid signature pattern analysis: residues are numbered based on the ORF1a/b absolute amino acid position (starting from number 1) in the reference sequence JQ065043 (Pig/China). The only residues with 100% frequency in Haitian strains mapped in the ORF1a/b and Spike regions. b) Spike glycoprotein trimer structure. Different colors are assigned to each monomer. Amino acid residues were numbered relative to their position in the Spike glycoprotein of the reference sequence. Red indicates residue 550, where Haitian sequences have a Valine to Alanine mutation. The other mutation (Proline to Leucine at position 38) was not resolved in the structure.