Tn-sequencing of Mycoplasma hyopneumoniae and Mycoplasma hyorhinis mutant libraries reveals non-essential genes of porcine mycoplasmas differing in pathogenicity

Bettina S Trueeb,Peter Kuhnert,Walid H Gharib,Simona Gerber,Dominiek Maes

doi:10.1186/s13567-019-0674-7

Bettina S Trueeb, Peter Kuhnert + Show 3 more

Open Access

https://doi.org/10.1186/s13567-019-0674-7

Copy DOI

Abstract

Mycoplasma hyopneumoniae and Mycoplasma hyorhinis are two phylogenetically related species colonizing the respiratory tract of pigs but differing in pathogenicity, the basis of which is not well resolved. We hypothesize that genes belonging to the species-specific portion of the genome and being non-essential during ideal laboratory growth conditions encode possible virulent determinants and are the driver of interspecies differences. To investigate this, transposon mutant libraries were generated for both species and a transposon sequencing (Tn-seq) method for mycoplasmas was established to identify non-essential genes. Tn-seq datasets combined with bidirectional Blastp analysis revealed that 101 out of a total 678 coding sequences (CDS) are species-specific and non-essential CDS of M. hyopneumoniae strain F7.2C, while 96 out of a total 751 CDS are species-specific and non-essential CDS in the M. hyorhinis strain JF5820. Among these species-specific and non-essential CDS were genes involved in metabolic pathways. In particular, the myo-inositol and the sialic acid pathways were found to be non-essential and therefore could be considered important to the specific pathogenicity of M. hyopneumoniae and M. hyorhinis, respectively. Such pathways could enable the use of an alternative energy source providing an advantage in their specific niche and might be interesting targets to knock out in order to generate attenuated live vaccines.

Highlights

Mycoplasmas form a genus of bacteria that has undergone reductive evolution from Gram-positive bacteria to wall-less cells with a small genome of high AT content
The species-specific set of genes for both Mycoplasma species was identified by bidirectional BLASTp analysis of their protein files and the non-essential genes of M. hyopneumoniae strain F7.2C and of M. hyorhinis strain JF5820 were investigated by sequencing corresponding transposon mutant libraries
The genome of M. hyorhinis strain JF5820 consisted of 840 423 bp with an average GC-content of 21.9%

Summary

Introduction

Mycoplasmas form a genus of bacteria that has undergone reductive evolution from Gram-positive bacteria to wall-less cells with a small genome of high AT content. They are highly host-adapted and often host-dependent [1]. To select relevant candidate genes of M. hyopneumoniae or M. hyorhinis and better explain their difference in pathogenicity, we aimed to identify the species-specific and non-essential genes of both species. We consider the species-specific portion of the genome of these phylogenetically related species being responsible for their difference in pathogenicity. The species-specific set of genes for both Mycoplasma species was identified by bidirectional BLASTp analysis of their protein files and the non-essential genes of M. hyopneumoniae strain F7.2C and of M. hyorhinis strain JF5820 were investigated by sequencing corresponding transposon mutant libraries

Objectives

Methods

Results

Conclusion