The efficacy and safety of closure of brachial access using the AngioSeal closure device: Experience with 161 interventions in diabetic patients with critical limb ischemia

PurposeThis study retrospectively evaluated the efficacy and safety of the 6F Angio-Seal (St. Jude Medical, St. Paul, Minn) as a closure device for transbrachial artery access for endovascular procedures in diabetic patients with critical limb ischemia.MethodsFrom January 2005 and September 2007, 1887 diabetic patients underwent interventional procedures in the lower limbs at a two diabetic foot centers. Patients presented with rest pain (16%), ulcers (80%), or gangrene (4%). Systemic anticoagulation with sodium heparin (70 IU/kg) was obtained for all patients at the beginning of the endovascular treatment. A total of 249 brachial arteries (238 patients) were evaluated for possible Angio-Seal use after endovascular recanalization of the leg. Color Doppler ultrasound imaging of the artery was obtained before revascularization only in patients with previous Angio-Seal placement in the brachial artery. No further imaging studies were done in the remaining brachial arteries where the Angio-Seal was deployed at the operator’s discretion. Impairment or disappearance of the radial pulse or onsets of hand ischemia or hand pain, or impairment of hand function during or at the end of the endovascular revascularization were all regarded as contraindications to Angio-Seal usage. Evidence of a highly calcified plaque of the brachial artery access site at the time of vessel puncture was regarded as an absolute contraindication to the Angio-Seal use. Patients were seen before discharge, at 1, 3, and 8 weeks after the procedure, and at 3-month intervals thereafter. Complications included hemorrhage, pseudoaneurysm, infection, and vessel occlusion.ResultsA total of 1947 Angio-Seal collagen plugs were deployed in 1709 diabetic patients (90.5%). The Angio-Seal was used for brachial artery closure in 159 patients (8.4%) in 161 procedures (159 in the left, 2 in the right brachial artery). In 79 patients (4.2%) in 88 procedures (87 in the left and 1 in the right brachial artery), the device was deemed contraindicated due to small vessel size in 73 patients (92.4%) or presence of calcium at the access site in five patients (6.3%). One patient (1.3%) refused the collagen plug closure after revascularization. The non-Angio-Seal group was evaluated for comparison. The success rate for achieving hemostasis in the Angio-Seal group was 96.9%. Five major complications (3.1%) at 30 days consisted of two puncture site hematomas >4 cm, two brachial artery occlusions, and one brachial artery pseudoaneurysm, with three patients requiring open surgery. Minor complications (7.50%) were three puncture site hematomas < 4 cm, three oozing of blood from the access site, and six patients had mild pain in the cubital fossa. No further complications were recorded in the 14-month follow-up (range 1-25 months) of a total of 140 patients.ConclusionsThis retrospective study shows that the 6F Angio-Seal is a valuable and safe vascular closure device for transbrachial access in diabetic patients undergoing interventional procedures for critical limb ischemia

Transient Hypermutagenesis Accelerates the Evolution of Legume Endosymbionts following Horizontal Gene Transfer

International audienceHorizontal gene transfer (HGT) is an important mode of adaptation and diversification of prokaryotes and eukaryotes and a major event underlying the emergence of bacterial pathogens and mutualists. Yet it remains unclear how complex phenotypic traits such as the ability to fix nitrogen with legumes have successfully spread over large phylogenetic distances. Here we show, using experimental evolution coupled with whole genome sequencing, that co-transfer of imuABC error-prone DNA polymerase genes with key symbiotic genes accelerates the evolution of a soil bacterium into a legume symbiont. Following introduction of the symbiotic plasmid of Cupriavidus taiwanensis, the Mimosa symbiont, into pathogenic Ralstonia solanacearum we challenged transconjugants to become Mimosa symbionts through serial plant-bacteria co-cultures. We demonstrate that a mutagenesis imuABC cassette encoded on the C. taiwanensis symbiotic plasmid triggered a transient hypermutability stage in R. solanacearum transconjugants that occurred before the cells entered the plant. The generated burst in genetic diversity accelerated symbiotic adaptation of the recipient genome under plant selection pressure, presumably by improving the exploration of the fitness landscape. Finally, we show that plasmid imuABC cassettes are over-represented in rhizobial lineages harboring symbiotic plasmids. Our findings shed light on a mechanism that may have facilitated the dissemination of symbiotic competency among a-and b-proteobacteria in natura and provide evidence for the positive role of environment-induced mutagenesis in the acquisition of a complex lifestyle trait. We speculate that co-transfer of complex phenotypic traits with mutagenesis determinants might frequently enhance the ecological success of HGT

Distribution of plasmid <i>nodABC</i> (p<i>nod</i>) and plasmid <i>imuBC</i> (p<i>imuBC</i>) genes among α- and β-proteobacteria.

<p>Blue and yellow rectangles indicate presence and absence of genes in the corresponding genome, respectively, as assayed by BlastP analysis. Dark blue rectangles indicate <i>nodABC</i> and <i>imuBC</i> genes co-localized on the same plasmid. α- and β-proteobacteria are arranged according to their position on the core genome phylogeny. Species of the same genus are similarly colored. See <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942.s009" target="_blank">Table S3</a> for details.</p

Experimental evolution of <i>R. solanacearum</i> into <i>Mimosa</i> symbionts.

<p>(A) <i>R. solanacearum</i> GMI1000 containing the <i>C. taiwanensis</i> symbiotic pRalta plasmid was evolved under <i>M. pudica</i> selection pressure. In a first step, three spontaneous <i>M. pudica</i>-nodulating derivatives of <i>GMI1000</i> pRalta, CBM212, CBM349, and CBM356 (selection cycle), were selected using <i>M. pudica</i> as a trap <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Marchetti1" target="_blank">[24]</a>. Nine independent lineages have been then derived from CBM212 (A–C), CBM349 (G–I), and CBM356 (M, N, S) using serial <i>M. pudica</i>-bacteria co-culture cycles of 21 days (evolution cycles) <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Marchetti1" target="_blank">[24]</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Guan1" target="_blank">[25]</a>. Green and red arrow heads indicate activation of nodulation and intracellular infection, respectively <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Marchetti1" target="_blank">[24]</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Guan1" target="_blank">[25]</a>. Numbers between brackets indicate the total number of point mutations detected in each clone relative to its closest re-sequenced ancestor. Point mutations are available on the Microscope platform (<a href="https://www.genoscope.cns.fr/agc/microscope/expdata/evoProject.php" target="_blank">https://www.genoscope.cns.fr/agc/microscope/expdata/evoProject.php</a>). (B–D) Nodulation and infection have been dramatically improved over 16 evolution cycles. <i>In planta</i> fitness (B) and nodulation competitiveness (C) of final clones relative to their respective nodulating ancestors, following equal co-inoculation of each of the nine final/ancestral pairs. Nodule infectiveness (D) of final clones (Ev) as compared to ancestors (An). Graphs summarize data from <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Guan1" target="_blank">[25]</a>,<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Marchetti2" target="_blank">[26]</a>. *<i>p</i>-value (t-test) <0.05. (E) In each cycle, bacteria were inoculated in the Jensen plant medium. Root nodules, which appeared from 5 days after inoculation, were each induced by a single bacterial cell that subsequently multiplied within nodule tissue <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942-Gage1" target="_blank">[56]</a>. In the selection and evolution cycles bacteria spent ∼21 days and from a few days up to 14 days in the plant medium, respectively. Population sizes are estimates. gen., generations. Raw data are provided in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942.s013" target="_blank">Data S1</a>.</p

Model for symbiotic and mutagenic plasmid-driven evolution of rhizobia.

<p>Following horizontal transfer of a symbiotic plasmid to a soil bacterium, the recipient genome accumulates environment-induced mutations that lead to phenotypic diversification. The most beneficial variants are selected by the plant and clonally multiply within nodules before being released. Rounds of <i>ex planta</i> phenotypic diversification/plant selection/clonal multiplication may have driven the adaptation process <i>in natura</i>.</p

Evolvability of <i>imuA2B2C2</i>⁺ populations.

<p>(A) Experimental evolution of <i>imuA2B2C2</i>⁺ and Δ<i>imuA2B2C2</i> nodulating chimeric <i>Ralstonia</i>. Each ancestor was evolved using serial <i>M.pudica</i>-bacteria co-culture cycles, either <i>ex planta-in planta</i> cycles of 21 days (nodule bacteria serving as inoculum in each cycle, red lines) or <i>ex planta</i> cycles of 7 days (rhizospheric bacteria serving as inoculum in each cycle, blue lines). For <i>ex planta</i> lineages, 7-day cycles were chosen since the mean time bacteria spent <i>ex planta</i> in the 16 cycle evolution experiment (<a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio-1001942-g001" target="_blank">Figure 1</a>) was estimated to be 7 days. Ancestors were antibiotic resistant derivatives of a <i>hrpG</i> mutant of GMI1000pRalta. Four to five independent lineages have been derived from each ancestor. SpeR, spectinomycin-resistant strain. KanR, kanamycin-resistant strain. (B) Relative <i>in planta</i> fitness of <i>imuA2B2C2</i>⁺ versus Δ<i>imuA2B2C2</i> populations following co-inoculations of final populations derived from <i>ex planta-in planta</i> lineages (red legend, all Ev-<i>imu</i>⁺(i) versus Ev-Δ<i>imu</i>(i) pairs) or from <i>ex planta</i> lineages (blue legend, all Ev-<i>imu</i>⁺(i′) versus Ev-Δ<i>imu</i>(i′) pairs). Nodule bacteria were counted 21 days after inoculation. *Indicates significant differences between ancestral <i>imu</i>⁺ clones and evolved <i>imu⁺</i> populations (t-test, <i>p</i><0.05). #Indicates significant differences between <i>Ev-imu</i>⁺(i) (evolved <i>ex planta-in planta</i>) and Ev-<i>imu⁺</i>(i′) (evolved <i>ex planta</i>) populations (t-test, <i>p</i><0.05). ‡Indicates significant differences between Ev-<i>imu</i>⁺ and Ev-Δ<i>imu</i> populations for each series of competition experiments (either Ev-<i>imu⁺</i>(i) versus Ev-Δ<i>imu</i>(i) or Ev-<i>imu⁺</i>(i′) versus Ev-Δ<i>imu</i>(i′), t-test, <i>p</i><0.001). See <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942.s005" target="_blank">Figure S5B and S5C</a> and <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942.s015" target="_blank">Data S3</a> for details. (C) Relative <i>ex planta</i> fitness of <i>imuA2B2C2</i>⁺ versus Δ<i>imuA2B2C2</i> populations following co-inoculations of final populations derived from <i>ex planta-in planta</i> lineages. Bacteria recovered from the Jensen medium were counted 7 days after inoculation in Gibson tubes containing <i>M. pudica</i> plants. <i>imuA2B2C2</i>⁺ ancestors better survived in Jensen-<i>Mimosa</i> than Δ<i>imuA2B2C2</i>, in accordance with results presented in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942.s002" target="_blank">Figure S2A</a>. ‡Indicates significant differences between <i>imu</i>⁺ and Δ<i>imu</i> populations for each series of competition experiments (either An-<i>imu</i>⁺(i) versus An-Δ<i>imu</i>(i) or Ev-<i>imu⁺</i>(i) versus Ev-Δ<i>imu</i>(i), t-test, <i>p</i><0.01). No significant difference was observed between ancestral <i>imu</i>⁺ clones and evolved <i>imu</i>⁺ populations. Raw data are provided in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001942#pbio.1001942.s015" target="_blank">Data S3</a>.</p