Intragenic tandem repeat variation between Legionella pneumophila strains

<p>Abstract</p> <p>Background</p> <p>Bacterial genomes harbour a large number of tandem repeats, yet the possible phenotypic effects of those found within the coding region of genes are only beginning to be examined. Evidence exists from other organisms that these repeats can be involved in the evolution of new genes, gene regulation, adaptation, resistance to environmental stresses, and avoidance of the immune system.</p> <p>Results</p> <p>In this study, we have investigated the presence and variability in copy number of intragenic tandemly repeated sequences in the genome of <it>Legionella pneumophila</it>, the etiological agent of a severe pneumonia known as Legionnaires' disease. Within the genome of the Philadelphia strain, we have identified 26 intragenic tandem repeat sequences using conservative selection criteria. Of these, seven were "polymorphic" in terms of repeat copy number between a large number of <it>L. pneumophila </it>serogroup 1 strains. These strains were collected from a wide variety of environments and patients in several geographical regions. Within this panel of strains, all but one of these seven genes exhibited statistically different patterns in repeat copy number between samples from different origins (environmental, clinical, and hot springs).</p> <p>Conclusion</p> <p>These results support the hypothesis that intragenic tandem repeats could play a role in virulence and adaptation to different environments. While tandem repeats are an increasingly popular focus of molecular typing studies in prokaryotes, including in <it>L. pneumophila</it>, this study is the first examining the difference in tandem repeat distribution as a function of clinical or environmental origin.</p

Functional MRI of Auditory Responses in the Zebra Finch Forebrain Reveals a Hierarchical Organisation Based on Signal Strength but Not Selectivity

BACKGROUND: Male songbirds learn their songs from an adult tutor when they are young. A network of brain nuclei known as the 'song system' is the likely neural substrate for sensorimotor learning and production of song, but the neural networks involved in processing the auditory feedback signals necessary for song learning and maintenance remain unknown. Determining which regions show preferential responsiveness to the bird's own song (BOS) is of great importance because neurons sensitive to self-generated vocalisations could mediate this auditory feedback process. Neurons in the song nuclei and in a secondary auditory area, the caudal medial mesopallium (CMM), show selective responses to the BOS. The aim of the present study is to investigate the emergence of BOS selectivity within the network of primary auditory sub-regions in the avian pallium. METHODS AND FINDINGS: Using blood oxygen level-dependent (BOLD) fMRI, we investigated neural responsiveness to natural and manipulated self-generated vocalisations and compared the selectivity for BOS and conspecific song in different sub-regions of the thalamo-recipient area Field L. Zebra finch males were exposed to conspecific song, BOS and to synthetic variations on BOS that differed in spectro-temporal and/or modulation phase structure. We found significant differences in the strength of BOLD responses between regions L2a, L2b and CMM, but no inter-stimuli differences within regions. In particular, we have shown that the overall signal strength to song and synthetic variations thereof was different within two sub-regions of Field L2: zone L2a was significantly more activated compared to the adjacent sub-region L2b. CONCLUSIONS: Based on our results we suggest that unlike nuclei in the song system, sub-regions in the primary auditory pallium do not show selectivity for the BOS, but appear to show different levels of activity with exposure to any sound according to their place in the auditory processing stream

Functional characterization of a Legionella pneumophila collagen-like protein encoded by a gene with a variable number of tandem repeats

Legionella pneumophila is een facultatief intracellulaire pathogee n die in de omgeving hoofdzakelijk in protozoa, maar ook vrijlevend en i n biofilms wordt terug gevonden. Infectie van de mens wordt veroorzaakt door inhalatie van gecontamineerde aerosols gevolgd door replicatie van de bacteriën in de alveolaire macrofagen en de longepitheelcellen. Deze infectie kan aanleiding geven tot een ernstige pneumonie, beter gekend a ls de veteranenziekte , of tot Pontiac koorts, een onschadelijke aandoe ning. Bacteriën bevatten vele repetitive sequenties, waaronder tandem repeats , een direkte opeenvolging van herhalingen. Sommige tandem repeats he bben een variabel aantal tandemherhalingen en dan definieert men deze r epeat regio als een VNTR ( variable number of tandem repeats ). VNTRs b ehoren tot de meest diverse loci in bacteriële populaties en worden freq uent geassocieerd met virulentie. Deze associatie met virulentie werd oo k aangetoond voor VNTRs in genen van L. pneumophila. Een nieuw gen (lpg 2644, lcl) met een VNTR-regio dat codeert voor een collageen-gelijkend eiwit (Lcl), werd geïdentificeerd in L. pneumophila. Het aantal tandemherhalingen in de VNTR-regio van < I&gt;lcl werd bestudeerd en de rol van het collageen-gelijkend eiwit, Lc l, in de virulentie van L. pneumophila werd onderzocht. Vooreerst werd, door middel van enzymatische experimenten, de aanwezighe id van de triple helix structuur, karakteristiek voor eukaryoot collagee n, in Lcl aangetoond. Virulentie-gerelateerde experimenten bevestigden d e rol van Lcl in de adhesie van L. pneumophila aan alveolaire macrofagen en longepitheelcellen en preliminaire resultaten suggereren bovendien dat de adhesiekarakteristieken van Lcl beïnvloed worden door h et aantal aanwezige tandemherhalingen in de VNTR-regio. Vervolgens werd de relatie tussen Lcl en twee collageen receptoren (inte grines en C1q-receptor) bestudeerd. L. pneumophila acitveerde het focaal adhesie kinase, een indicator voor integrine binding, maar e en specifieke interactie tussen integrines en Lcl werd niet aangetoond. De complement C1q-receptor, daarentegen, die affiniteit heeft voor de co llageensequentie van de complement factor C1q en de longcollectines, pro fileerde zich wel als een directe interactiepartner van Lcl. Daar vasthechting een eerste stap is voor intracellulaire replicatie en deze vasthechting deels gemedieerd wordt door Lcl, mogen we besluiten da t Lcl inderdaad een rol speelt in virulentie van L. pneumophila< /&gt;. De VNTR-regio van lcl werd in meer detail bestudeerd in een diverse collectie van 108 L. pneumophila serotype I-stamm en. De studie van het aantal tandemherhalingen resulteerde in de identif icatie van 12 polymorfismen, gesitueerd tussen 7 en 19 tandemherhalingen . Voor de klinische en omgevingsisolaten was er geen correlatie tussen h et aantal tandemherhalingen en de oorsprong van de stam waar te nemen. O pvallend was wel dat er slechts 3 polymorfismen terug te vinden waren in de stammen geïsoleerd uit bronnen met een hogere temperatuur. Deze obse rvaties suggereren dat het aantal tandemherhalingen geen belangrijke rol speelt in humane infecties, maar van belang is in overleving van < I&gt;L. pneumophila bij hoge temperatuur. Tot slot is de geobserveerde v ariatie in het aantal tandemherhalingen een resultaat van verschillende enzymen, waaronder RecA. RecA is een multifunctioneel enzyme dat een bel angrijke rol speelt in bacteriële DNA recombinatie en DNA herstel. In de ze studie werd aangetoond dat RecA deels verantwoordelijk was voor het v ariabel aantal tandemherhalingen en dat de invloed van RecA afhankelijk was van het aantal tandemherhalingen.nrpages: 137status: publishe

Role of the Halloween genes, Spook and Phantom in ecdysteroidogenesis in the desert locust, Schistocerca gregaria

The functional characterization of the Halloween genes represented a major breakthrough in the elucidation of the ecdysteroid biosynthetic pathway. These genes encode cytochrome P450 enzymes catalyzing the final steps of ecdysteroid biosynthesis in the dipteran Drosophila melanogaster and the Lepidoptera Manduca sexta and Bombyx mori. This is the first report on the identification of two Halloween genes, spook (spo) and phantom (phm), from a hemimetabolous orthopteran insect, the desert locust Schistocerca gregaria. Using q-RT-PCR, their spatial and temporal transcript profiles were analyzed in both final larval stage and adult locusts. The circulating ecdysteroid titers in the hemolymph were measured and found to correlate well with changes in the temporal transcript profiles of spo and phm. Moreover, an RNA interference (RNAi)-based approach was employed to study knockdown effects upon silencing of both transcripts in the fifth larval stage. Circulating ecdysteroid levels were found to be significantly reduced upon dsRNA treatment.status: publishe

Evaluating an easy sampling method using dried blood spots to determine vedolizumab concentrations

An association between vedolizumab (VDZ) trough concentrations and therapeutic outcome has been observed in patients with inflammatory bowel diseases. VDZ samples are typically collected via venous sampling for therapeutic drug monitoring (TDM), but can alternatively be collected via dried blood spot (DBS) samples, which can be used for intensive sampling to investigate pharmacokinetic profiles. Therefore, we have developed a DBS method for determining VDZ concentrations and validated this method by comparing VDZ measurements in paired DBS and venous patient samples. First, VDZ was spiked in citrated whole blood and spotted on filter paper. After drying, DBS were extracted and VDZ concentrations were determined in the extracts using ELISA. For clinical validation, 41 paired DBS and serum samples were collected from 19 VDZ-treated patients. VDZ concentrations measured in DBS extracts strongly correlated with serum concentrations (Pearson r = 0.978, p < 0.0001). No significant impact of the hematocrit value was observed on the VDZ DBS/serum concentration ratios. Additionally, the VDZ DBS/serum ratio was calculated in nine individual patients, which was, in general, shown to be stable over time. VDZ DBS sampling is a robust method that can be used as an alternative to venous blood collection for TDM of VDZ. VDZ concentrations in DBS highly correlated with VDZ serum concentrations over a broad concentration range, allowing DBS to be used for intensive sampling to gain more insight in VDZ pharmacokinetics.status: publishe

Neuropeptide Receptors as Possible Targets for Development of Insect Pest Control Agents

Vaious insect species have a severe impact on human welfare and environment and thus force us to continuously develop novel agents for pest control. Neuropeptides constitute a very versatile class of bioactive messenger molecules that initiate and/or regulate a wide array of vital biological processes in insects by acting on their respective receptors in the plasmamembrane of target cells. These receptors belong to two distinct categories of signal transducing proteins, i.e., heptahelical or G protein-coupled receptors (7TM, GPCR) and single transmembrane containing receptors. An increasing amount ofevidence indicates that insect neuropeptide-receptor couples play crucial roles in processes as diverse as development, metabolism, ecdysis and reproduction. As such, they gain growing interest as promising candidate targets for the development of a new generation of species- and receptor-specific insect control agents that may generate fewer side effects. In this chapter, we will present some examples of insect neuropeptide receptors and aim to demonstrate their fundamental importance in insect biology.Advances in Experimental Medicine and Biologystatus: publishe

Comparative genomics of leucine-rich repeats containing G protein-coupled receptors and their ligands.

Leucine-rich repeats containing G protein-coupled receptors (LGRs) constitute a unique cluster of transmembrane proteins sharing a large leucine-rich extracellular domain for hormone binding. In mammals, LGRs steer important developmental, metabolic and reproductive processes as receptors for glycoprotein hormones and insulin/relaxin-related proteins. In insects, a receptor structurally related to human LGRs mediates the activity of the neurohormone bursicon thereby regulating wing expansion behaviour and remodelling of the newly synthesized exoskeleton. In the past decade, novel insights into the molecular evolution of LGR encoding genes accumulated rapidly due to comparative genome analyses indicating that the endocrine LGR signalling system likely emerged before the radiation of metazoan phyla and expanded throughout evolution. Here, we present a short survey on the evolution of LGRs and the hormones they interact with.Comparative StudyJournal ArticleResearch Support, Non-U.S. Gov'tReviewSCOPUS: sh.jinfo:eu-repo/semantics/publishe

Purification and characterization of an insulin-related peptide in the desert locust, Schistocerca gregaria: immunolocalization, cDNA cloning, transcript profiling and interaction with neuroparsin

Members of the insulin superfamily are not restricted to vertebrates, but have also been identified in invertebrate species. In the current report, we present the characterization of Scg-insulin-related peptide (IRP), an insulin-related peptide in the desert locust, Schistocerca gregaria. This peptide was isolated from corpora cardiaca (CC) extracts by means of a high-performance liquid chromatography (HPLC)-based purification strategy. Subsequent cloning and sequencing of the corresponding cDNA revealed that the encoded Scg-IRP precursor displays the structural organization that is typical for members of the insulin superfamily. Moreover, immunocytochemistry on brain tissue sections demonstrated the presence of Scg-IRP in median neurosecretory cells of the pars intercerebralis and their projections towards the storage part of the CC. Quantitative real-time RT-PCR studies revealed the presence of Scg-IRP transcripts in a variety of tissues, including nervous tissue and fat body. Furthermore, these transcripts showed a tissue- and phase-dependent, temporal regulation during the reproductive cycle of adult males and females. Finally, we demonstrated that Scg-IRP interacts in vitro with a recombinant neuroparsin, a locust protein displaying sequence similarity with vertebrate IGF binding proteins.status: publishe