Biology subjects

Tram, G.

Publications and source records attributed to Tram, G..

4 recordsLinked to original sources

bioRxiv2022

Actinobacillus pleuropneumoniae encodes multiple phase-variable DNA methyltransferases that comprise distinct phasevarions

ABSTRACTActinobacillus pleuropneumoniae is the cause of porcine pleuropneumonia, a severe respiratory tract infection that is responsible for major economic losses to the swine industry. Many host-adapted bacterial pathogens encode systems known as phasevarions (phase- variable regulons). Phasevarions result from variable expression of cytoplasmic DNA methyltransferases. Variable expression results in genome-wide methylation differences within a bacterial population, leading to altered expression of multiple genes via epigenetic mechanisms. Our examination of a diverse population of A. pleuropneumoniae strains determined that Type I and Type III DNA methyltransferases with the hallmarks of phase variation were present in this species. We demonstrate that phase variation is occurring in these methyltransferase, and show associations between particular Type III methyltransferase alleles and serovar. Using Pacific BioSciences Single-Molecule, Real-Time (SMRT) sequencing and Oxford Nanopore sequencing, we demonstrate the presence of the first ever characterised phase-variable, cytosine-specific Type III DNA methyltransferase. Phase variation of distinct Type III DNA methyltransferase variants results in the regulation of distinct phasevarions, and in multiple phenotypic differences relevant to pathobiology. Our characterisation of these newly described phasevarions in A. pleuropneumoniae will aid in the selection of stably expressed antigens, and direct and inform development of a rationally designed subunit vaccine against this major veterinary pathogen.

microbiology↗

bioRxiv2021

Characterisation of the phase-variable autotransporter Lav reveals a role in host cell adherence and biofilm formation in Non-Typeable Haemophilus influenzae

Lav is an autotransporter protein found in pathogenic Haemophilus and Neisseria species. Lav in non-typeable Haemophilus influenzae (NTHi) is phase-variable: the gene reversibly switches ON-OFF via changes in length of a locus-located GCAA(n) simple DNA sequence repeat tract. The expression status of lav was examined in carriage and invasive collections of NTHi, where it was predominantly not expressed (OFF). Phenotypic study showed lav expression (ON) results in increased adherence to host cells, and denser biofilm formation. A survey of Haemophilus spp. genome sequences showed lav is present in [~]60% of NTHi strains, but lav is not present in most typeable H. influenzae. Sequence analysis revealed a total of five distinct variants of the Lav passenger domain present in Haemophilus spp., with these five variants showing a distinct lineage distribution. Determining the role of Lav in NTHi will help understand the role of this protein during distinct pathologies.

microbiology↗

bioRxiv2021

Streptococcus suis encodes multiple allelic variants of a phase-variable Type III DNA methyltransferase, ModS, that control distinct phasevarions

Streptococcus suis is a significant cause of bacterial meningitis in humans, particularly in S.E. Asia, and is a leading cause of respiratory and invasive disease in pigs. Phase-variable DNA methyltransferases, associated with Restriction-Modification (R-M) systems, are a source of epigenetic gene regulation, controlling the expression of multiple genes. These systems are known as phasevarions (phase-variable regulons), and have been characterised in many host-adapted bacterial pathogens. We recently described the presence of a Type III DNA methyltransferase in S. suis, ModS, which contains a simple sequence repeat (SSR) tract within the open reading frame of the modS gene, and which varied in length between individual strains. We also observed multiple allelic variants of the modS gene were present in a population of S. suis isolates. Here, we demonstrate that a biphasic ON-OFF switching of expression occurs in the two most common ModS alleles, ModS1 and ModS2, and that switching is dependent on SSR tract length. Further, we show that ModS1 and ModS2 are active methyltransferases in S. suis using Single-Molecule, Real Time (SMRT) sequencing. ON-OFF switching of each ModS allele results in the regulation of distinct phasevarions, with the ModS2 phasevarion impacting growth patterns and antibiotic resistance. This is the first demonstration of a phase-variable Type III DNA methyltransferase in a Gram-positive organism that controls a phasevarion. Characterising the phenotypic effects of phasevarions in S. suis is key to understanding pathogenesis and the development of future vaccines. ImportanceStreptococcus suis is a causative agent of meningitis, polyarthritis and polyserositis in swine, and is a major cause of zoonotic meningitis in humans. Here we investigate epigenetic gene regulation in S. suis by multiple phasevarions controlled by the phase-variable Type III DNA methyltransferase ModS. This is the first characterised example of a Type III R-M system regulating a phasevarion in a Gram-positive organism. We demonstrate that biphasic ON-OFF switching of ModS expression results in differences in bacterial growth and antibiotic resistance. Understanding the effects of ModS phase variation is required to determine the stably expressed antigenic repertoire of S. suis, which will direct and inform the development of antimicrobial treatments and vaccines against this important pathogen.

microbiology↗

bioRxiv2019

Assigning a role for chemosensory signal transduction in Campylobacter jejuni biofilms using a combined omics approach

Biofilms serve as a protective mechanism for bacteria to cope with environmental stress. Whilst ordinarily a fastidious organism, it has been long suggested that C. jejuni is able to utilise this mode of growth as a way to transmit infection from the avian host to humans. Herein, we undertook a combinatorial approach to examine differential expression of C. jejuni genes and protein abundance during biofilm formation. RNA sequencing and proteomics via quantitative liquid chromatography - tandem mass spectrometry (LC-MS/MS) revealed biofilm growth induced a substantial rearrangement of the C. jejuni transcriptome and proteome, with [~]600 genes differentially expressed in biofilms compared to planktonic cells. Biofilm-induced genes / proteins were those involved in iron metabolism and acquisition, cell division, glycan production and attachment, while those repressed were associated with metabolism, amino acid uptake and utilisation, and large tracts of the chemotaxis pathway. We further examined the role of chemotaxis in C. jejuni biofilm formation by assessing the behaviour of isogenic strains with deletions of the cheV and cheW genes. Both {Delta}cheV and {Delta}cheW exhibited a significant decrease in directed motility when compared to wild-type C. jejuni. Both mutants also demonstrated an increase in autoagglutination ability and increased biofilm formation. A subtle difference was also observed between the phenotypes of {Delta}cheV and {Delta}cheW mutants, both in motility and biofilm formation. This suggests roles for the CheV and CheW signal transduction proteins and may present signal transduction as a potential method for modulating C. jejuni biofilm formation. Author summaryCampylobacter jejuni is a gastroenteric bacterium that is responsible for most cases of bacterial food poisoning in the developed world. The organism commonly resides in avian reservoirs and is passed to humans through contaminated poultry and animal products. Ordinarily, C. jejuni requires a strict set of conditions in order to survive and cause infections in humans. Biofilms are a method of bacterial growth that may provide shielding from harsh environments and provide an important link between reservoirs and human hosts. In this study, we have utilised a multi-platform approach to compare gene expression and protein abundance in planktonic C. jejuni cells and those growing as a biofilm. We subsequently focused on the chemosensory system of C. jejuni and demonstrated that signal transduction proteins play a role in biofilm formation. Our work has provided a broad profile of which genes are important to C. jejuni biofilms and that the chemosensory pathway has an influence on biofilm formation.

microbiology↗