Genetic Organisation, Mobility and Predicted Functions of Genes on Integrated, Mobile Genetic Elements in Sequenced Strains of Clostridium difficile

Abstract

Background Clostridium difficile is the leading cause of hospital-associated diarrhoea in the US and Europe. Recently the incidence of C. difficile-associated disease has risen dramatically and concomitantly with the emergence of ‘hypervirulent’ strains associated with more severe disease and increased mortality. C. difficile contains numerous mobile genetic elements, resulting in the potential for a highly plastic genome. In the first sequenced strain, 630, there is one proven conjugative transposon (CTn), Tn5397, and six putative CTns (CTn1, CTn2 and CTn4-7), of which, CTn4 and CTn5 were capable of excision. In the second sequenced strain, R20291, two further CTns were described.ResultsCTn1, CTn2 CTn4, CTn5 and CTn7 were shown to excise from the genome of strain 630 and transfer to strain CD37. A putative CTn from R20291, misleadingly termed a phage island previously, was shown to excise and to contain three putative mobilisable transposons, one of which was capable of excision. In silico probing of C. difficile genome sequences with recombinase gene fragments identified new putative conjugative and mobilisable transposons related to the elements in strains 630 and R20291. CTn5-like elements were described occupying different insertion sites in different strains, CTn1-like elements that have lost the ability to excise in some ribotype 027 strains were described and one strain was shown to contain CTn5-like and CTn7-like elements arranged in tandem. Additionally, using bioinformatics, we updated previous gene annotations and predicted novel functions for the accessory gene products on these new elements.ConclusionsThe genomes of the C. difficile strains examined contain highly related CTns suggesting recent horizontal gene transfer. Several elements were capable of excision and conjugative transfer. The presence of antibiotic resistance genes and genes predicted to promote adaptation to the intestinal environment suggests that CTns play a role in the interaction of C. difficile with its human host.

Diversity and Evolution in the Genome of Clostridium difficile

“…Some C. difficile transposons (Tns) are mobilizable, meaning that they rely on complex host-mediated mechanisms for conjugation and full mobility (111,112). Some elements are self-transmissible and are known as conjugative transposons (CTns) or integrative and conjugative elements (ICEs).…”

“…The introduction of these elements into the C. difficile genome leads to numerous heritable changes not only in the acquisition of new, possibly advantageous genes but also, as in the case of insertion within an open reading frame (ORF), in gene disruption and phenotypic alteration (112,114). Dissemination of these elements and their accessory genes by lateral transfer has significantly contributed to the genetic diversity seen in C. difficile and possibly contributed to the success of C. difficile as an opportunistic pathogen (52,73,112,114,115).…”

“…Dissemination of these elements and their accessory genes by lateral transfer has significantly contributed to the genetic diversity seen in C. difficile and possibly contributed to the success of C. difficile as an opportunistic pathogen (52,73,112,114,115).…”

“…Unlike these and other related pathogens, resistance to antimicrobials in C. difficile is unusual in that it is mediated predominantly by Tns as opposed to plasmids (52,112,114). Through traditional PCR-and, more recently, WGS-based studies, numerous Tns associated with antimicrobial resistance in C. difficile have been described, including (120), and Tn5398.…”

Behavior and Target Site Selection of Conjugative Transposon Tn 916 in Two Different Strains of Toxigenic Clostridium difficile

“…A second, not mutually exclusive hypothesis is that these host genomes evolve to lose target sites which are present within open reading frames (ORFs) in order to protect their genes. The fact that C. difficile has such a large amount of intergenic DNA may be one of the reasons for the large number of integrative elements that are present within this host (5,30). It appears that target site selection is an interaction of element and host properties.…”