Integrative Conjugative Elements Are Widespread in Field Isolates of Mycoplasma Species Pathogenic for Ruminants

Abstract

Comparative genomics have revealed massive horizontal gene transfer (HGT) between Mycoplasma species sharing common ruminant hosts. Further results pointed toward an integrative conjugative element (ICE) as an important contributor of HGT in the small-ruminant-pathogen Mycoplasma agalactiae. To estimate the prevalence of ICEs in ruminant mycoplasmas, we surveyed their occurrence in a collection of 166 field strains representing 4 (sub)species that are recognized as major pathogens. Based on available sequenced genomes, we first defined the conserved, minimal ICE backbone as composed of 4 coding sequences (CDSs) that are evenly distributed and predicted to be essential for ICE chromosomal integration-excision and horizontal transfer. Screening of the strain collection revealed that these 4 CDSs are well represented in ruminant Mycoplasma species, suggesting widespread occurrence of ICEs. Yet their prevalence varies within and among species, with no correlation found with the individual strain history. Extrachromosomal ICE forms were also often detected, suggesting that ICEs are able to circularize in all species, a first and essential step in ICE horizontal transfer. Examination of the junction of the circular forms and comparative sequence analysis of conserved CDSs clearly pointed toward two types of ICE, the hominis and spiroplasma types, most likely differing in their mechanism of excision-integration. Overall, our data indicate the occurrence and maintenance of functional ICEs in a large number of field isolates of ruminant mycoplasmas. These may contribute to genome plasticity and gene exchanges and, presumably, to the emergence of diverse genotypes within pathogenic mycoplasmas of veterinary importance.M ycoplasma species represent a large group of wall-less bacteria belonging to the class Mollicutes that have often been portrayed as minimal cells because of the small size of their genomes and the paucity of their metabolic pathways. In the tree of life reconstructed from 31 conserved proteins (1), mycoplasmas are depicted on some of the longest branches, suggesting that they have evolved quickly. Indeed, a recent coalescent analysis supports this hypothesis by estimating the nucleotide substitution rate in Mycoplasma gallisepticum to be one of the highest reported for bacteria (2). Regarding the content of genes, it has long been considered that successive losses of genetic material were the only driving force of mycoplasma evolution (3), a scenario in agreement with these minimal bacteria having adopted a parasitic lifestyle. However, recent in silico genomic analyses revealed that mycoplasmas sharing the same habitat have exchanged a significant amount of their genome via horizontal gene transfer (HGT) (4). This was particularly true for the ruminant Mycoplasma species that are distributed into two distinct phylogenetic groups but share common animal hosts and ecological niches (e.g., the ear canal, the udder, the respiratory tract, etc.), with some having exchanged up to 18% of their genomes (5). Rumin...

The Integrative Conjugative Element (ICE) ofMycoplasma agalactiae: key elements involved in horizontal dissemination and influence of co-resident ICEs

“…The minimal ICEA chassis was consistent with the conservation of cds5, cds17, cds19 and cds22 across documented ICEs of ruminant mycoplasma species (8), and the occurrence of cds1, cds14 and cds16 at very similar location in a majority of MICEs (2, 4–6, 23). Interestingly, the conjugative properties of M. agalactiae were also abrogated by mTn insertion in NCRs raising questions regarding the presence of regulatory elements and/or key motifs, such as an oriT , in these regions.…”

“…Since their discovery in mycoplasma species of the Hominis phylogenetic group, MICEs have been found broadly distributed across Mollicutes (2–6, 8) and their pivotal role in HGTs is emerging (7, 12, 13). Taking advantage of the M. agalactiae ICE prototype, this study provides the first functional analysis of MICE factors involved in conjugative transfer.…”

“…These self-transmissible elements encode their excision, transfer by conjugation and integration into the genome of the recipient cell where they replicate as a part of the host chromosome. Recently, a new family of self-transmissible integrative elements has been identified in the genome of several mycoplasma species that confers conjugative properties to this important group of bacteria (2–8).…”

Loss of diversity within Mycoplasma bovis isolates collected in France from bovines with respiratory diseases over the last 35 years

“…bovis is well-equipped to generate diversity. Indeed sequenced genomes have revealed insertion sequences, integrative and conjugative elements (ICEs) and vsp loci (Li et al, 2011;Qi et al, 2012;Wise et al, 2011) and field strains have recently been shown to often harbor functional ICEs (Tardy et al, 2014). As a logical consequence, the genetic heterogeneity of M. bovis isolates collected at different times has seemed to be the rule in many countries, and was reported in Europe (Kusiluka et al, 2000;McAuliffe et al, 2004;Sulyok et al, 2014), North America (BellRogers et al, 2012;Soehnlen et al, 2011) and Israel (Amram et al, 2013) etc.…”

Removal of a subset of non-essential genes fully attenuates a highly virulentMycoplasmastrain

“…The lipoproteins encoded in the D3 (MMCAP2_0014-0016; 4,677-bp region; D3) as well as the D5 region ( lppQ , MMCAP2_0889-0904; 24,906-bp region, D5) were also excised employing again the yeast-based engineering method. Finally, we deleted a large genomic region that contained the Mycoplasma -specific F1-likeX0 ATPase (15), the MIB-MIP system (9) that has been shown in vitro to modulate the action of immunoglobulins through specific degradation, an integrative and conjugative element (ICE) (16) and several genes encoding other lipoproteins. The ICE was targeted in an effort to reduce mobile elements from the Mmc genome.…”