Analysis of the Global Population Structure of Paenibacillus larvae and Outbreak Investigation of American Foulbrood Using a Stable wgMLST Scheme.

Bojan Papić,Margo Diricks,Darja Kušar

doi:10.3389/fvets.2021.582677

Abstract

Paenibacillus larvae causes the American foulbrood (AFB), a highly contagious and devastating disease of honeybees. Whole-genome sequencing (WGS) has been increasingly used in bacterial pathogen typing, but rarely applied to study the epidemiology of P. larvae. To this end, we used 125 P. larvae genomes representative of a species-wide diversity to construct a stable whole-genome multilocus sequence typing (wgMLST) scheme consisting of 5745 loci. A total of 51 P. larvae isolates originating from AFB outbreaks in Slovenia were used to assess the epidemiological applicability of the developed wgMLST scheme. In addition, wgMLST was compared with the core-genome MLST (cgMLST) and whole-genome single nucleotide polymorphism (wgSNP) analyses. All three approaches successfully identified clusters of outbreak-associated strains, which were clearly separated from the epidemiologically unlinked isolates. High levels of backward comparability of WGS-based analyses with conventional typing methods (ERIC-PCR and MLST) were revealed; however, both conventional methods lacked sufficient discriminatory power to separate the outbreak clusters. The developed wgMLST scheme provides an improved understanding of the intra- and inter-outbreak genetic diversity of P. larvae and represents an important progress in unraveling the genomic epidemiology of this important honeybee pathogen.

Highlights

Paenibacillus larvae is a Gram-positive bacterium that is the causative agent of American foulbrood (AFB), a devastating disease affecting honeybee (Apis mellifera) larvae [1]
The final whole-genome multilocus sequence typing (wgMLST) scheme consisted of 5738 wgMLST loci and seven loci from the conventional multilocus sequence typing (MLST) scheme described previously [6] and is commercially available through a plugin in BioNumerics
We applied Whole-genome sequencing (WGS) to study the molecular epidemiology of P. larvae on a set of 179 genomes representative of a species-wide diversity

Summary

Introduction

Paenibacillus larvae is a Gram-positive bacterium that is the causative agent of American foulbrood (AFB), a devastating disease affecting honeybee (Apis mellifera) larvae [1]. Several genotyping methods have been applied to study the molecular epidemiology and population structure of P. larvae, including (i) the repetitive element PCR fingerprinting (rep-PCR), in particular the enterobacterial repetitive intergenic consensusbased polymerase chain reaction (ERIC-PCR) [4], (ii) pulsed-field gel electrophoresis (PFGE) [5], and (iii) multilocus sequence typing (MLST) [6]. These conventional methods give roughly concordant results with regard to P. larvae distribution (i.e., clearly separate the epidemiologically most relevant types ERIC I and II), but with various degree of discriminatory power and repeatability [5, 6]. ERIC types differ in their phenotypic characteristics, including their virulence [5, 7,8,9]

Methods

Results

Conclusion