First Animal Source Metagenome Assembly of Lawsonella clevelandensis from Canine External Otitis

Abstract

External otitis is one of the most common conditions in dogs to be presented to the veterinarian. Moreover, the disorder is often challenging to manage. The range and role of microorganisms involved in the pathogenesis are currently not fully understood. Therefore, the condition has been studied using third-generation sequencing (Oxford Nanopore Technology) to gain a more complete picture of the pathogens involved. Throughout the metagenome assembly of a sample from the ear canal of an 11-year-old female Yorkshire terrier suffering from chronic external otitis, a genome of Lawsonella clevelandensis was compiled. To our knowledge, this result is the first of its type of animal origin. The outcome of the assembly is a single circular chromosome with a length of 1,909,339 bp and 1727 predicted genes. One open reading frame associated with antimicrobial resistance could have been identified. Comparing all available genomes, the species can be associated with three main genome clusters. The finding contributes to the extending knowledge bank about this often-overlooked pathogen and raises attention to the role of nanopore sequencing by the identification and characterization of microorganisms that are difficult to culture.

Keywords: Lawsonella clevelandensis; dog; metagenome-assembled genome; nanopore sequencing; otitis externa.

Figure 1

Pan-genome structure of Lawsonella clevelandensis strains. (A) The number of gene families in each gene set. The categories indicate the proportion of genomes in which a gene family is present: Core genes (90–100%), shell genes (15% to <89%), and cloud genes (0% to <15%). (B) Gene accumulation curves for the pan-genome. (C) Gene presence–absence heatmap visualizing the distribution of gene families across genomes. Each row corresponds to a gene family, and each column represents an individual strain. Red and blue indicate gene presence and absence, respectively. Both rows and columns are hierarchically clustered based on Euclidean distance.

Figure 2

COG functional annotation. (A) Percentage rates of COG functional categories associated with the L. clevelandensis assembly. (B) Comparison of the L. clevelandensis pan-genome and the studied genome. The COG categories are represented by the same colors in plots (A,B).

Figure 3

Gene-tree based on transaldolase amino acid sequences. RefSeq assembly IDs are shown, and the Nocardiopsis dassonvillei (GCF_900638215) was used as an outgroup. Numbers at branches indicate bootstrap support levels (100 replicates). The assemblies originate from abscesses (GCF_001281505, GCF_001293125, GCF_900610365), thorax (GCF_040117505), hospital NICU environment (GCF_003241315), human gut (GCF_032574585), oral cavity (GCF_905373635), and skin metagenomes (GCF_030826395, GCF_043427535, GCF_936933995, GCF_943913485, GCF_946223255).