Molecular Characteristics, Ecology, and Zoonotic Potential of Escherichia coli Strains That Cause Hemorrhagic Pneumonia in Animals

Abstract

Hemorrhagic pneumonia (HP) is a rare but highly lethal disease, mainly of dogs and cats, caused by hemolytic Escherichia coli strains that contain cnf1 (encoding cytotoxic necrotizing factor 1). After encountering fatal HP in two dogs, we used contemporary molecular methods, including multilocus sequence typing and whole-genome sequencing, to compare the corresponding case isolates with published HP clinical isolates and newly obtained fecal E. coli isolates from 20 humans and animals in the index HP case household. We also compared the aggregated HP clinical isolates, which represented 13 discrete strains, by pulsotype with a large, private pulsotype library of diverse-source E. coli. The HP clinical isolates represented a narrow range of phylogenetic group B2 lineages (mainly sequence types 12 and 127), O types (mainly O4 and O6), and H types (mainly H5 and H31), but diverse fimH alleles (type-1 fimbriae adhesin). Their extensive, highly conserved virulence genotypes, which qualified as extraintestinal pathogenic E. coli (ExPEC), encoded diverse adhesins, toxins, iron uptake systems, and protectins. Household surveillance identified multiple HP-like fecal strains, plus abundant between-host strain sharing, including of the household's index HP strain. The pulsotype library search identified, for five HP clinical strains, same-pulsotype human and animal fecal and clinical (predominantly urine) isolates, from diverse locales and time periods. Thus, E. coli strains that cause HP derive from a narrow range of ExPEC lineages within phylogroup B2, contain multiple virulence genes other than cnf1, are shared extensively between hosts, and likely function in nature mainly as intestinal colonizers and uropathogens. IMPORTANCE This study clarifies the clonal background and extensive virulence genotypes of the E. coli strains that cause hemorrhagic pneumonia in domestic animals (mainly dogs and cats), shows that such strains circulate among animals and humans, identifies a substantial intestinal colonization component to their lifestyle, and extends their known clinical manifestations to include bacteremia and urinary tract infection. The findings place these strains better into context vis-à-vis current understandings of E. coli phylogeny, ecology, and pathogenesis; identify questions for future research; and may prove relevant for surveillance and prevention efforts.

Keywords: Escherichia coli; ExPEC; cytotoxic necrotizing factor; hemorrhagic pneumonia; microbial ecology; molecular epidemiology; phylogenetic analysis; transmission; virulence determinants; whole-genome sequencing.

FIG 1

Light and electron microscopy of lung tissue from case 1 of hemorrhagic pneumonia due to Escherichia coli. (Panel 1) (Hematoxylin and eosin [H&E] stain) Near-total effacement of normal architecture by hemorrhage, edema, fibrin, and necrotic debris. Bar, 200 μm. (Panel 2) (H&E stain) Alveolar walls are effaced. Alveolar spaces are filled with hemorrhage, edema, fibrin, and pyknotic mononuclear cells. Bar, 100 μm. (Panel 3) (Gram stain) Gram-negative bacilli (arrows) aggregate within alveolar spaces abutting the pleura. Bar, 50 μm. (Panel 4) (Transmission electron micrograph [TEM]) Alveolar space (star) contains erythrocytes (asterisks) and alveolar capillaries (black arrows). Capillary lumen (CL) contains fibrin thrombus. White arrow, pyknotic capillary endothelial cell. Bar, 5 μm. (Panel 5) (TEM) Alveolar wall (arrows) is expanded by fibrin strands and cell debris. Bar, 2 μm. (Panel 6) (TEM) Macrophage with bacteria in a phagolysosome (black arrows). Bar, 1 μm.

FIG 2

Core-genome phylogram for 13 Escherichia coli isolates from animals with hemorrhagic pneumonia. The (unrooted) phylogram was inferred according to the maximum-likelihood method by using SNP Tree in Enterobase, based on 89,170 variant sites. Isolate 0.2025 (ST127–O6:H31) was the reference genome. Only variant sites present in all 13 isolates were included. Isolates JJ2691 and JJ3030 are from cases 1 and 2, respectively, presented here. Other isolates are archival (previously published) isolates.