The Role of Minor Pilins in Assembly and Function of the Competence Pilus of Streptococcus pneumoniae

The remarkable genomic plasticity of Streptococcus pneumoniae largely depends on its ability to undergo natural genetic transformation. To take up extracellular DNA, S. pneumoniae assembles competence pili composed of the major pilin ComGC. In addition to ComGC, four minor pilins ComGD, E, F, and G are expressed during bacterial competence, but their role in pilus biogenesis and transformation is unknown. Here, using a combination of protein-protein interaction assays we show that all four proteins can directly interact with each other. Pneumococcal ComGG stabilizes the minor pilin ComGD and ComGF and can interact with and stabilize the major pilin ComGC, thus, deletion of ComGG abolishes competence pilus assembly. We further demonstrate that minor pilins are present in sheared pili fractions and find ComGF to be incorporated along the competence pilus by immunofluorescence and electron microscopy. Finally, mutants of the invariant Glu5 residue (E5), ComGDE5A or ComGEE5A, but not ComGFE5A, were severely impaired in pilus formation and function. Together, our results suggest that ComGG, lacking E5, is essential for competence pilus assembly and function, and plays a central role in connecting the pneumococcal minor pilins to ComGC.

Ribosomal protein S1 plays a critical role in horizontal gene transfer by mediating the expression of foreign mRNAs

The emergence of multi-antibiotic resistant bacteria is one of the largest threats to global heath. This rise is due to the genomic plasticity of bacteria, allowing rapid acquisition of antibiotic resistance through the uptake of foreign DNA (i.e. horizontal gene transfer, HGT). This genomic plasticity is not limited to DNA from bacteria, highly divergent (trans-kingdom) mRNA have been reported to drive translation in E. coli. Trans-kingdom activity has been attributed to mRNA tertiary structure suggesting the bacterial translation machinery bottle-necks HGT, restricting the expression of foreign DNA. However, here we show that tertiary structure is not responsible for ribosome recruitment and that the translation efficiency is dependent on ribosomal protein S1 and an A-rich Shine-Dalgarno-like element. The S1-facilitated ability of ribosomes to identify and exploit A-rich sequences in foreign RNA highlights the important role that S1 plays in horizontal gene transfer, the robustness of canonical prokaryotic translation, and bacterial evolution.

The genomes of nucleocytoplasmic large DNA viruses: viral evolution writ large

The nucleocytoplasmic large DNA viruses (NCLDVs) are a diverse group that currently contain the largest known virions and genomes, also called giant viruses. The first giant virus was isolated and described nearly 20 years ago. Their genome sizes were larger than for any other known virus at the time and it contained a number of genes that had not been previously described in any virus. The origin and evolution of these unusually complex viruses has been puzzling, and various mechanisms have been put forward to explain how some NCLDVs could have reached genome sizes and coding capacity overlapping with those of cellular microbes. Here we critically discuss the evidence and arguments on this topic. We have also updated and systematically reanalysed protein families of the NCLDVs to further study their origin and evolution. Our analyses further highlight the small number of widely shared genes and extreme genomic plasticity among NCLDVs that are shaped via combinations of gene duplications, deletions, lateral gene transfers and de novo creation of protein-coding genes. The dramatic expansions of the genome size and protein-coding gene capacity characteristic of some NCLDVs is now increasingly understood to be driven by environmental factors rather than reflecting relationships to an ancient common ancestor among a hypothetical cellular lineage. Thus, the evolution of NCLDVs is writ large viral, and their origin, like all other viral lineages, remains unknown.

Genome-wide characterization of Mariner Like transposons and their derived MITEs in the Whitefly Bemisia tabaci (Hemiptera: Aleyrodidae)

The whitefly, Bemisia tabaci is a hemipteran pest of vegetable crops vectoring a broad category of viruses. Currently, this insect pest showed a high adaptability and resistance to almost all the chemical compounds commonly used for its control. In many cases, Transposable Elements (TEs) contributed to the evolution of host genomic plasticity. This study focuses on the annotation of Mariner Like Elements (MLEs) and their derived Miniature Inverted repeat Transposable Elements (MITEs) in the genome of B. tabaci. Two full-length MLEs belonging to mauritiana and irritans subfamilies were detected and named Btmar1.1 and Btmar2.1, respectively. Additionally, 548 defective MLE sequences clustering mainly into 19 different Mariner lineages of mauritiana and irritans subfamilies were identified. Each subfamily showed a significant variation in MLE copy number and size. Furthermore, 71 MITEs were identified as MLEs derivatives that could be mobilized via the potentially active transposases encoded by Btmar 1.1 and Btmar2.1. The vast majority of sequences detected in the whitefly genome present unusual Terminal Inverted Repeats (TIRs) of up to 400 bp in length. However, some exceptions are sequences without TIRs. This feature of the MLEs and their derived MITEs in B. tabaci genome that distinguishes them from all the other MLEs so far described in insects, which have TIRs size ranging from 20 to 40 bp. Overall, our study provides an overview of MLEs, especially those with large TIRs, and their related MITEs, as well as diversity of their families, which will provide a better understanding of the evolution and adaptation of the whitefly genome.

Identification of two variants of Acinetobacter baumannii 17978 with distinct genotypes and phenotypes

Acinetobacter baumannii is a nosocomial pathogen that exhibits substantial genomic plasticity. Here, the identification of two variants of A. baumannii ATCC 17978 that differ based on the presence of a 44 kb accessory locus, which was named AbaAL44 (“ A. baumannii accessory locus 44 kb”), is described. Analyses of existing deposited data suggest that both variants are found in published studies of A. baumannii ATCC 17978 and that ATCC-derived laboratory stocks are comprised of a mix of these two variants. Yet, each variant exhibits distinct interactions with the host in vitro and in vivo . Infection with the variant that harbors AbaAL44 ( Ab 17978 UN) results in decreased bacterial burdens and increased neutrophilic lung inflammation in a mouse model of pneumonia, and affects the production of IL-1β and IL-10 by infected macrophages. AbaAL44 harbors putative pathogenesis genes including those predicted to encode for a type I pilus cluster, a catalase, and a cardiolipin synthase. The accessory catalase increases A. baumannii resistance to oxidative stress and neutrophil-mediated killing in vitro . The accessory cardiolipin synthase plays a dichotomous role by promoting bacterial uptake and increasing IL-1β production by macrophages, but also enhancing bacterial resistance to cell envelope stress. Collectively, these findings highlight the phenotypic consequences of A. baumannii’s genomic dynamism through the evolution of two variants of a common type strain with distinct infection-related attributes.

Genetic Characterisation of Colistin Resistant Klebsiella pneumoniae Clinical Isolates From North India

BackgroundIncreasing use of colistin has led to the world-wide emergence of mobile colistin resistant gene (mcr). The present study aimed to identify and characterise mcr and other drug-resistant genes in colistin resistant Klebsiella pneumoniae clinical isolates.MethodsTwenty-two colistin resistant K. pneumoniae were analysed for mcr and other drug-resistant genes, efflux pumps, and virulence genes, and for their biofilm forming ability. Pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) were performed for all mcr-1 positive isolates. S1-PFGE and Southern hybridisation were performed for localisation of mcr-1 and blaNDM.ResultsNineteen colistin resistant K. pneumoniae harboured mcr-1 and 3 had mgrB disruption. All isolates harboured blaOXA-48-type and ESBL genes; eight strains (five with mcr-1 and three with mgrB disruption) co-harboured blaNDM. Efflux pumps genes AcrAB and mdtK were detected in all 22 and tol-C in 21 isolates. Virulence-related genes entB and irp-1 were detected in all 22, mrkD in 20, and fimH-1 in 18 isolates; 11 isolates were strong biofilm producers. PFGE clustered mcr-1 positive isolates into eight groups based on ≥90% similarity; MLST revealed diverse sequence types, predominant being ST-15 (n = 4) and ST-16 (n = 4). Both mcr-1 and blaNDM were localised on plasmid and chromosome; mcr-1 was present on IncFII type and blaNDM on IncFIB and IncA/C type plasmids.ConclusionsColistin resistance in K. pneumoniae was predominantly mediated by mcr-1. Co-existence of colistin, carbapenem, and other drug-resistant genes along with efflux pumps indicates towards enormous genomic plasticity in K. pneumoniae with ability to emerge as super-spreader of drug-resistance.

Diversity and evolution of pigment types and the phycobilisome rod gene region of marine Synechococcus cyanobacteria

Synechococcus picocyanobacteria are ubiquitous and abundant photosynthetic organisms in the marine environment and contribute for an estimated 16% of the ocean net primary productivity. Their light-harvesting complexes, called phycobilisomes (PBS), are composed of a conserved allophycocyanin core from which radiates six to eight rods with variable phycobiliprotein and chromophore content. This variability allows Synechococcus to optimally exploit the wide variety of spectral niches existing in marine ecosystems. Seven distinct pigment types or subtypes have been identified so far in this taxon, based on the phycobiliprotein composition and/or the proportion of the different chromophores in PBS rods. Most genes involved in their biosynthesis and regulation are located in a dedicated genomic region called the PBS rod region. Here, we examined the variability of gene sequences and organization of this genomic region in a large set of sequenced isolates and natural populations of Synechococcus representative of all known pigment types. All regions start with a tRNA-PheGAA and some possess mobile elements including tyrosine recombinases, suggesting that their genomic plasticity relies on a tycheposon-like mechanism. Comparison of the phylogenies obtained for PBS and core genes revealed that the evolutionary history of PBS rod genes differs from the rest of the genome and is characterized by the co-existence of different alleles and frequent allelic exchange. We propose a scenario for the evolution of the different pigment types and highlight the importance of population-scale mechanisms in maintaining a wide diversity of pigment types in different Synechococcus lineages despite multiple speciation events.

Underground metabolism facilitates the evolution of novel pathways for vitamin B6 biosynthesis

The term vitamin B6 is a designation for the vitamers pyridoxal, pyridoxamine, pyridoxine and the respective phosphate esters pyridoxal-5′-phosphate (PLP), pyridoxamine-5′-phosphate and pyridoxine-5′-phosphate. Animals and humans are unable to synthesise vitamin B6. These organisms have to take up vitamin B6 with their diet. Therefore, vitamin B6 is of commercial interest as a food additive and for applications in the pharmaceutical industry. As yet, two naturally occurring routes for de novo synthesis of PLP are known. Both routes have been genetically engineered to obtain bacteria overproducing vitamin B6. Still, major genetic engineering efforts using the existing pathways are required for developing fermentation processes that could outcompete the chemical synthesis of vitamin B6. Recent suppressor screens using mutants of the Gram-negative and Gram-positive model bacteria Escherichia coli and Bacillus subtilis, respectively, carrying mutations in the native pathways or heterologous genes uncovered novel routes for PLP biosynthesis. These pathways consist of promiscuous enzymes and enzymes that are already involved in vitamin B6 biosynthesis. Thus, E. coli and B. subtilis contain multiple promiscuous enzymes causing a so-called underground metabolism allowing the bacteria to bypass disrupted vitamin B6 biosynthetic pathways. The suppressor screens also show the genomic plasticity of the bacteria to suppress a genetic lesion. We discuss the potential of the serendipitous pathways to serve as a starting point for the development of bacteria overproducing vitamin B6. Key points • Known vitamin B6 routes have been genetically engineered. • Underground metabolism facilitates the emergence of novel vitamin B6 biosynthetic pathways. • These pathways may be suitable to engineer bacteria overproducing vitamin B6.

Genomic plasticity and antibody response of Bordetella bronchiseptica strain HT200, a natural variant from a thermal spring

ABSTRACT Classical Bordetella species are primarily isolated from animals and humans causing asymptomatic infection to lethal pneumonia. However, isolation of these bacteria from any extra-host environmental niche has not been reported so far. Here, we have characterized the genomic plasticity and antibody response of Bordetella bronchiseptica strain HT200, isolated from a thermal spring. Genomic ANI value and SNPs-based phylogenetic tree suggest a divergent evolution of strain HT200 from a human-adapted lineage of B. bronchiseptica. Growth and survivability assay showed strain HT200 retained viability for more than 5 weeks in the filter-sterilized spring water. In addition, genes or loci encoding the Bordetella virulence factors such as DNT, ACT and LPS O-antigen were absent in strain HT200, while genes encoding other virulence factors were highly divergent. Phenotypically, strain HT200 was non-hemolytic and showed weak hemagglutination activity, but was able to colonize in the respiratory organs of mice. Further, both infection and vaccination with strain HT200 induced protective antibody response in mouse against challenge infection with virulent B. bronchiseptica strain RB50. In addition, genome of strain HT200 (DSM 26023) showed presence of accessory genes and operons encoding predicted metabolic functions pertinent to the ecological conditions of the thermal spring.

Pangenomics of the Symbiotic Rhizobiales. Core and Accessory Functions Across a Group Endowed with High Levels of Genomic Plasticity

Pangenome analyses reveal major clues on evolutionary instances and critical genome core conservation. The order Rhizobiales encompasses several families with rather disparate ecological attitudes. Among them, Rhizobiaceae, Bradyrhizobiaceae, Phyllobacteriacreae and Xanthobacteriaceae, include members proficient in mutualistic symbioses with plants based on the bacterial conversion of N2 into ammonia (nitrogen-fixation). The pangenome of 12 nitrogen-fixing plant symbionts of the Rhizobiales was analyzed yielding total 37,364 loci, with a core genome constituting 700 genes. The percentage of core genes averaged 10.2% over single genomes, and between 5% to 7% were found to be plasmid-associated. The comparison between a representative reference genome and the core genome subset, showed the core genome highly enriched in genes for macromolecule metabolism, ribosomal constituents and overall translation machinery, while membrane/periplasm-associated genes, and transport domains resulted under-represented. The analysis of protein functions revealed that between 1.7% and 4.9% of core proteins could putatively have different functions.