Genetic Analysis of the Capsular Biosynthetic Locus from All 90 Pneumococcal Serotypes

Several major invasive bacterial pathogens are encapsulated. Expression of a polysaccharide capsule is essential for survival in the blood, and thus for virulence, but also is a target for host antibodies and the basis for effective vaccines. Encapsulated species typically exhibit antigenic variation and express one of a number of immunochemically distinct capsular polysaccharides that define serotypes. We provide the sequences of the capsular biosynthetic genes of all 90 serotypes of Streptococcus pneumoniae and relate these to the known polysaccharide structures and patterns of immunological reactivity of typing sera, thereby providing the most complete understanding of the genetics and origins of bacterial polysaccharide diversity, laying the foundations for molecular serotyping. This is the first time, to our knowledge, that a complete repertoire of capsular biosynthetic genes has been available, enabling a holistic analysis of a bacterial polysaccharide biosynthesis system. Remarkably, the total size of alternative coding DNA at this one locus exceeds 1.8 Mbp, almost equivalent to the entire S. pneumoniae chromosomal complement

La pena de treballs en benefici de la comunitat : prevenció i gestió de les incidències en el seu compliment

Aquest estudi té com a objectiu explorar la prevenció i gestió d'incidències que duen a terme els delegats d'execució de mesures per tal d'aconseguir un compliment efectiu, per part dels penats a TBC. Així, a través del marc teòric es vol abordar el compliment en general per tal de comprendre la seva importància des de diferents perspectives així com els elements que la literatura científica mostra com a dificultats en el procés de compliment. El treball de camp recull les aportacions de delegades d'execució de mesures mitjançant una metodologia qualitativa.This study aims to explore the prevention and management of incidents carried out by probation officers aimed at the effective compliance, by those sentenced to community service. Thus, the theoretical framework aims to address compliance in general in order to understand its importance from different perspectives as well as the elements that the scientific literature shows as difficulties in the compliance process. The fieldwork collects the visions of probation officers using a qualitative methodology

Protocol: genetic transformation of the fern Ceratopteris richardii through microparticle bombardment

BACKGROUND: The inability to genetically transform any fern species has been a major technical barrier to unlocking fern biology. Initial attempts to overcome this limitation were based on transient transformation approaches or achieved very low efficiencies. A highly efficient method of stable transformation was recently reported using the fern Ceratopteris richardii, in which particle bombardment of callus tissue achieved transformation efficiencies of up to 72%. As such, this transformation method represents a highly desirable research tool for groups wishing to undertake fern genetic analysis. RESULTS: We detail an updated and optimized protocol for transformation of C. richardii by particle bombardment, including all necessary ancillary protocols for successful growth and propagation of this species in a laboratory environment. The C. richardii lifecycle comprises separate, free-living gametophyte and sporophyte stages. Callus is induced from the sporophyte apex through growth on cytokinin-containing tissue culture medium and can be maintained indefinitely by sub-culturing. Transgene DNA is introduced into callus cells through particle bombardment, and stable genomic integration events are selected by regeneration and growth of T(0) sporophytes for a period of 8 weeks on medium containing antibiotics. Selection of T(1) transgenic progeny is accomplished through screening T(1) gametophytes for antibiotic resistance. In many cases sexual reproduction and development of transgenic embryos requires growth and fertilization of gametophytes in the absence of antibiotics, followed by a separate screen for antibiotic resistance in the resultant sporophyte generation. CONCLUSIONS: Genetic transformation of C. richardii using this protocol was found to be robust under a broad range of bombardment and recovery conditions. The successful expansion of the selection toolkit to include a second antibiotic for resistance screening (G-418) and different resistance marker promoters increases the scope of transformations possible using this technique and offers the prospect of more complex analysis, for example the creation of lines carrying more than one transgene. The introduction of a robust and practicable transformation technique is a very important milestone in the field of fern biology, and its successful implementation in C. richardii paves the way for adoption of this species as the first fern genetic model. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13007-015-0080-8) contains supplementary material, which is available to authorized users

Complete Genome Sequence of Uropathogenic Proteus mirabilis, a Master of both Adherence and Motility▿ †

The gram-negative enteric bacterium Proteus mirabilis is a frequent cause of urinary tract infections in individuals with long-term indwelling catheters or with complicated urinary tracts (e.g., due to spinal cord injury or anatomic abnormality). P. mirabilis bacteriuria may lead to acute pyelonephritis, fever, and bacteremia. Most notoriously, this pathogen uses urease to catalyze the formation of kidney and bladder stones or to encrust or obstruct indwelling urinary catheters. Here we report the complete genome sequence of P. mirabilis HI4320, a representative strain cultured in our laboratory from the urine of a nursing home patient with a long-term (≥30 days) indwelling urinary catheter. The genome is 4.063 Mb long and has a G+C content of 38.88%. There is a single plasmid consisting of 36,289 nucleotides. Annotation of the genome identified 3,685 coding sequences and seven rRNA loci. Analysis of the sequence confirmed the presence of previously identified virulence determinants, as well as a contiguous 54-kb flagellar regulon and 17 types of fimbriae. Genes encoding a potential type III secretion system were identified on a low-G+C-content genomic island containing 24 intact genes that appear to encode all components necessary to assemble a type III secretion system needle complex. In addition, the P. mirabilis HI4320 genome possesses four tandem copies of the zapE metalloprotease gene, genes encoding six putative autotransporters, an extension of the atf fimbrial operon to six genes, including an mrpJ homolog, and genes encoding at least five iron uptake mechanisms, two potential type IV secretion systems, and 16 two-component regulators

Capsule Biosynthesis Genes and Repeat-Unit Polysaccharide Structures

<p>Shown are the <i>cps</i> gene clusters for cases discussed in the text, together with the polysaccharide structure of the encoded repeat unit where known [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0020031#pgen-0020031-b031" target="_blank">31</a>] (the full set is shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0020031#pgen-0020031-sg001" target="_blank">Figure S1</a>). Genes are represented on the forward and reverse strands by boxes coloured according to the gene key, with gene designations indicated above each box. Grey blocks indicate regions of sequence similarity between gene clusters. Repeat-unit structures are displayed with the linkage to undecaprenyl pyrophosphate at the right-hand side (not necessarily the case for the published structures [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0020031#pgen-0020031-b031" target="_blank">31</a>]), so residue numbers are counted from right to left. Monosaccharides are represented as shapes coloured according to the structure key. Housekeeping sugars are coloured grey. Non-housekeeping sugar colours correspond to the associated sugar biosynthesis gene colours. Glycerol, choline, and acetate are indicated as text. Also shown are the nature of linkages with the associated gene, and the linkages between repeat units created by the Wzy polymerase. Gene designations are in parentheses where their substrate specificity is unclear.</p