The genome and transcriptome of Haemonchus contortus, a key model parasite for drug and vaccine discovery

<p>Background: The small ruminant parasite Haemonchus contortus is the most widely used parasitic nematode in drug discovery, vaccine development and anthelmintic resistance research. Its remarkable propensity to develop resistance threatens the viability of the sheep industry in many regions of the world and provides a cautionary example of the effect of mass drug administration to control parasitic nematodes. Its phylogenetic position makes it particularly well placed for comparison with the free-living nematode Caenorhabditis elegans and the most economically important parasites of livestock and humans.</p> <p>Results: Here we report the detailed analysis of a draft genome assembly and extensive transcriptomic dataset for H. contortus. This represents the first genome to be published for a strongylid nematode and the most extensive transcriptomic dataset for any parasitic nematode reported to date. We show a general pattern of conservation of genome structure and gene content between H. contortus and C. elegans, but also a dramatic expansion of important parasite gene families. We identify genes involved in parasite-specific pathways such as blood feeding, neurological function, and drug metabolism. In particular, we describe complete gene repertoires for known drug target families, providing the most comprehensive understanding yet of the action of several important anthelmintics. Also, we identify a set of genes enriched in the parasitic stages of the lifecycle and the parasite gut that provide a rich source of vaccine and drug target candidates.</p> <p>Conclusions: The H. contortus genome and transcriptome provides an essential platform for postgenomic research in this and other important strongylid parasites. </p&gt

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

The OPERA trial : protocol for a randomised trial of an exercise intervention for older people in residential and nursing accommodation

Background Depression is common in residents of Residential and Nursing homes (RNHs). It is usually undetected and often undertreated. Depression is associated with poor outcomes including increased morbidity and mortality. Exercise has potential to improve depression, and has been shown in existing trials to improve outcomes among younger and older people. Existing evidence comes from trials that are short, underpowered and not from RNH settings. The aim of the OPERA trial is to establish whether exercise is effective in reducing the prevalence of depression among older RNH residents. Method OPERA is a cluster randomised controlled trial. RNHs are randomised to one of two groups with interventions lasting 12 months Intervention group: a depression awareness and physical activity training session for care home staff, plus a whole home physical activation programme including twice weekly physiotherapist-led exercise groups. The intervention lasts for one year from randomisation, or Control group: a depression awareness training session for care home staff. Participants are people aged 65 or over who are free of severe cognitive impairment and willing to participate in the study. Our primary outcome is the prevalence of depressive symptoms, a GDS-15 score of five or more, in all participants at the end of the one year intervention period. Our secondary depression outcomes include remission of depressive symptoms and change in GDS-15 scores in those with depressive symptoms prior to randomisation. Other secondary outcomes include, fear of falling, mobility, fractures, pain, cognition, costs and health related quality of life. We aimed to randomise 77 RNHs. Discussion Home recruitment was completed in May 2010; 78 homes have been randomised. Follow up will finish in May 2011 and results will be available late 2011

Untersuchungen zur Bedeutung des präpartalen Progesteronentzugs in Hinblick auf die Steuerung der Geburt und die Ablösung der Nachgeburt beim Rind

Beim Rind ist der präpartale Abfall der maternalen Progesteron (P4)-Konzentration Voraussetzung für den Eintritt der Geburt. Der P4-Abfall spiegelt beim Rind primär den Funktionsverlust des Trächtigkeitsgelbkörpers wider. Über die Bedeutung der plazentaren P4-Produktion lagen bisher praktisch keine Informa¬tionen vor. Die Rinder¬plazenta trägt in der Endphase der Gravidität zwar nur minimal zu den maternalen P4-Spiegeln bei, jedoch bildet sie hohe lokale P4-Konzentrationen im Bereich der feto-maternalen Kontakt¬zone. Diese könnten, vermittelt über P4-Rezeptoren (PR) in den maternalen Karunkeln, ein wesentlicher Faktor für die Differenzie¬rung und Funktion der Plazentome sein. Es wurde die Arbeits¬hypothese ent¬wickelt, dass der Entzug hoher lokaler P4-Konzentrationen ein wesentliches Signal für die Vorbereitung eines termin¬gerechten Nachgeburtsabganges darstellt. Entsprechend könnte ein unvollständiges Sistieren der plazentaren P4-Produktion vor der Geburt eine wesentliche Ursache für das Auftreten idiopathischer Nachge-burts¬verhaltungen sein. Zur Überprüfung dieser Hypo¬these wurden drei gravide Kühe am 270. und 271. Graviditätstag mit dem Antigestagen Aglepriston (Ap) behandelt (Gruppe D272+Ap). Dieser PR-Blocker ermöglicht die Ausschaltung aller rezeptor¬vermittelten P4-Wirkungen unabhängig von der P4-Quelle. Als Kontrollen dienten Tiere mit termin¬gerechter spontaner Geburt und termingerechtem Nachgeburts¬abgang (Gruppe Normalgeburt, n = 4, Trächtigkeitsdauer: 280,5 ± 1,7 Tage) sowie unbehan¬delte Tiere mit Schnitt¬entbindung am Tag 272 (Gruppe D272-Ap, n = 3). Die Kühe wurden klinisch überwacht und der Geburtsverlauf dokumentiert. Von allen Tieren wurden Blutproben in regelmäßigen Abständen sowie Plazentome unmittelbar im Anschluss an die Geburt (Gruppen D272+Ap und Normalgeburt) bzw. während der Schnittentbindungen (Gruppe D272-Ap) entnommen. In den Blutproben wurden die Konzentrationen von Progesteron und Östrogenen mittels radioimmuno¬logischer Verfahren und die 13, 14-Di¬hydro-15-Keto-PGF2α (PGFM)-Konzentrationen mittels ELISA gemessen. Als Parameter für die präpartale Umstrukturierung der Plazentome wurde der prozen¬tuale Anteil der Trophoblast-riesenzellen (TGC) an den Trophoblastzellen und die Reduktion des Karunkelepithels erfasst. Weiterhin wurde die Expression von Cyclooxygenase II (Cox II), PR und Glucocorticoidrezeptor (GR) auf Protein- und mRNA-Ebene beurteilt. Die Aglepristonbehandlung führte bei allen drei Kühen zu einer vorzeitigen Terminie¬rung der Gravidität. Erste Geburtsanzeichen traten 46,3 ± 6,0 Stunden nach Be¬handlungsbeginn auf. Es kam zur vollständigen Öffnung der Zervix, eine adäquate Wehentätigkeit setzte jedoch innerhalb der folgenden zwei Stunden nicht ein. So wurde ein manueller Auszug der Kälber durchgeführt. Neben der Öffnung der Zervix wurde durch die Antigestagenbehandlung die Laktogenese induziert. Entge¬gen der eigenen Hypothese wiesen alle drei Kühe der Gruppe D272+Ap, wie die Tiere der Gruppe D272-Ap, eine komplette Nachgeburts¬ver¬haltung auf und die Kälber beider Gruppen waren gleichermaßen prämatur. Die histo¬morphologischen Unter¬suchungen bestätigten, dass durch die Aglepriston¬behand¬lung die präpartale „Plazentareifung“ nicht induziert wurde. So waren, im Gegensatz zur Normalgeburtsgruppe, in den unreifen Plazentomen der Gruppen D272+Ap und D272−Ap weder ein Rück¬gang des relativen Anteils der TGC noch eine Reduktion des Karunkelepithels nach¬weisbar. Überraschenderweise wurde durch die Antigestagenbehandlung die Luteolyse induziert, erkennbar an einem steilen Abfall der P4-Werte vor bzw. während des Auszugs der Kälber. Korrespondierend mit dem Abfall der P4-Konzen¬tra¬tionen wurde bei den Tieren der Gruppe D272+Ap bereits präpartal ein schwacher Anstieg der PGFM-Werte beobachtet. Da bei diesen Tieren, im Gegensatz zur Normal¬geburtsgruppe, keine Aufregulation der plazentaren Cox II-Expression nach¬weisbar war, ist anzunehmen, dass die Antigestagen-induzierte Luteolyse indirekt durch Prostaglandine extraplazentaren, vermutlich endometrialen, Ursprungs aus¬gelöst wurde. Zum Zeitpunkt des Auszugs waren die PGFM-Plasma¬konzentrationen im Vergleich zur Normalgeburtsgruppe jedoch relativ gering (2,14 ± 1,40 ng/ml vs. 8,70 ± 2,20 ng/ml). Somit erklärt vermutlich ein Mangel an uterotonem PGF2α die Wehen¬schwäche bei den Aglepriston-behandelten Tieren. Die Östrogen¬synthese im Trophoblasten sowie die GR- bzw. PR-Expression in den Plazentomen wurden durch das Antigestagen nicht beeinflusst. Insgesamt lassen die Ergebnisse dieser Arbeit darauf schließen, dass nur ein relativ geringer Anteil der geburtsassoziierten Veränderungen direkt durch den präpartalen Progesteronentzug ausgelöst wird, nämlich die Öffnung der Zervix und das Einsetzen der Laktation. Dagegen erfordern andere wesentliche geburtsassoziierte Vorgänge, wie eine adäquate Wehentätigkeit und die Ablö¬sung der Plazenta, offensichtlich primär Signale aus dem fetalen Kompartiment. Die eigenständige Bedeutung der plazentaren P4¬-Produktion bleibt unklar.In cattle the prepartal decline in maternal progesterone (P4) levels is a prerequisite for the onset of parturition. This P4 withdrawal predominantly reflects the loss of luteal function, and virtually no information on the importance of placental P4 production is available. Despite its minimal contribution to maternal P4 levels in late gestation the bovine placenta is capable of producing high P4 levels locally at the feto-maternal interface, which – mediated by progesterone receptors (PR) previously detected in the maternal caruncles – could be an essential factor in placental differentiation and function. Thus, the hypothesis was put forward that a well-timed and complete withdrawal of high local P4 concentrations is a crucial signal for the timely release of the placenta. According to this concept, an incomplete cessation of placental P4 production during the initiation of parturition could be an important factor in the etiology of placental retention. To test for this hypothesis, three cows were treated with the antiprogestin aglepristone (Ap) on days 270 and 271 of gestation to abolish receptor mediated effects of P4 irrespective of its origin (group D272+Ap). As controls, four cows giving spontaneous birth at normal term (280.5 ± 1.7 days, group NT) with timely release of fetal membranes and three cows undergoing cesarean section on day 272 (group D272-Ap) were included into the study. The cows were monitored clinically, and the progress of birth was registered. From all animals, blood samples were taken regularly during the experimental period, and placentomes were removed per vaginam immediately after birth (groups D272+Ap and NT) or during cesarean section (group D272-Ap). In blood samples, con-centrations of P4 and estrogens were measured by radioimmunological methods, and for the measurement of 13, 14-dihydro-15-keto prostaglandin F2α (PGFM)-concentrations a commercial ELISA kit was used. To characterize the prepartal remodeling of placentomal microarchitecture, the percentage of trophoblast giant cells (TGC) relative to the total number of trophoblast cells and the reduction of caruncular epithelium were determined. Moreover, the expression of cyclo-oxygenase II (Cox II), PR and glucocorticoid receptor (GR) was investigated at protein and mRNA level. The application of aglepristone significantly reduced gestational length. First signs of impending parturition occurred 46.3 ± 6.0 hours after the start of treatment, and vaginal exploration confirmed that the cervix was fully open during this time. However, no progress in the expulsion of the calves could be observed during the following two hours, obviously due to insufficient myometrial activity. Consequently the calves were extracted. Besides a complete opening of the cervix, antiprogestin treatment induced the onset of lactation. Inconsistent with the working hypothesis, in all cows of the treatment group a severe retention of fetal membranes was observed. Similar cases of retained fetal membrane were also observed in all D272-Ap cows, and calves of both groups were slightly premature to a similar extent. Consistent with clinical observations histological investigations of placentomes showed that antiprogestin treatment did not induce placental maturation, whereas for the placentomes of NT cows the prepartal decline in TGC numbers and the reduction of caruncular epithelium was confirmed. Surprisingly, antiprogestin treatment induced luteolysis, as a significant decline of progesterone concentrations started before or during the expulsion of the calf. Con¬comitant with the decline in P4 concentrations in D272+Ap cows an increase of PGFM levels became detectable. The fact that, different from NT animals, no up-regu¬lation of placental Cox II was found suggests that the antiprogestin induced luteolysis indirectly stimulated prostaglandin pro¬duction from an extraplacental source, presumably the intercaruncular endometrium. However, at parturition PGFM levels in D272+Ap cows were clearly lower in comparison to NT cows (2.14 ± 1.40 ng/ml vs. 8.70 ± 2.20 ng/ml), which suggests that insufficient myometrial activity observed in antiprogestin treated cows was related to a reduced availability of uterotonic PGF2α. Placental estrogen production and the expression of PR or GR were not affected by antiprogestin treatment. In conclusion, the results demonstrate that only a minor part of the processes related to bovine parturition is directly dependent on P4 withdrawal, in particular the opening of the cervix and the onset of lactation. Moreover, they suggest that other important processes such as adequate myometrial activity and timely release of the placenta are predominantly dependent on signals from the fetal compartment. The importance of placental P4 production in cattle remains unknown

Attachment and invasion of Neisseria meningitidis to host cells is related to surface hydrophobicity, bacterial cell size and capsule

We compared exemplar strains from two hypervirulent clonal complexes, strain NMB-CDC from ST-8/11 cc and strain MC58 from ST-32/269 cc, in host cell attachment and invasion. Strain NMB-CDC attached to and invaded host cells at a significantly greater frequency than strain MC58. Type IV pili retained the primary role for initial attachment to host cells for both isolates regardless of pilin class and glycosylation pattern. In strain MC58, the serogroup B capsule was the major inhibitory determinant affecting both bacterial attachment to and invasion of host cells. Removal of terminal sialylation of lipooligosaccharide (LOS) in the presence of capsule did not influence rates of attachment or invasion for strain MC58. However, removal of either serogroup B capsule or LOS sialylation in strain NMB-CDC increased bacterial attachment to host cells to the same extent. Although the level of inhibition of attachment by capsule was different between these strains, the regulation of the capsule synthesis locus by the two-component response regulator MisR, and the level of surface capsule determined by flow cytometry were not significantly different. However, the diplococci of strain NMB-CDC were shown to have a 1.89-fold greater surface area than strain MC58 by flow cytometry. It was proposed that the increase in surface area without changing the amount of anchored glycolipid capsule in the outer membrane would result in a sparser capsule and increase surface hydrophobicity. Strain NMB-CDC was shown to be more hydrophobic than strain MC58 using hydrophobicity interaction chromatography and microbial adhesion-to-solvents assays. In conclusion, improved levels of adherence of strain NMB-CDC to cell lines was associated with increased bacterial cell surface and surface hydrophobicity. This study shows that there is diversity in bacterial cell surface area and surface hydrophobicity within N. meningitidis which influence steps in meningococcal pathogenesis

Neisserial LptA::His_x6 transfers PEA to lipid A of <i>E. coli</i> LPS.

<p>Lipid A profiles of LPS extracted from <i>E. coli</i> strains JCB571 expressing <i>Ec</i>DsbA (CKEC272) (Panel A), <i>E. coli</i> JCB571 expressing LptA::His_x6 (CKEC543) (Panel B) and JCB571 expressing LptA::His_x6 and <i>Ec</i>DsbA (CKEC564) (Panel C) as determined by MALDI-TOF MS. <i>bis</i>-Phosphorylated hexaacylated lipid A (m/z = 1796), the mono-phosphorylated derivative (<i>m/z</i> = 1716), and the heptaacylated version due to the addition of a palmitic acyl residue (<i>m/z</i> = 2034) were detected in all strains. <i>bis</i>-Phosphorylated tetraacylated lipid A (<i>m/z</i> = 1360) was found abundantly in the MALDI spectra of all three strains, which was likely produced from <i>bis</i>-phosphorylated hexaacylated lipid A (<i>m/z</i> = 1796) during the ionization step on MALDI. The lipid A preparations from CKEC543 expressing LptA (Panel B) and CKEC564 co-expressing LptA and <i>Ec</i>DsbA (Panel C) also contained ions consistent with one PEA added to the <i>bis</i>-phosphorylated structure (such as <i>m/z</i> 1919; i.e. 1796+123) and the heptaacylated structure (such as <i>m/z</i> = 2157, i.e. 2034+123).</p

Plasmids used in this study.

<p>*Cassettes: <i>aadA</i>  =  spectinomycin resistance, <i>tetM</i>  =  tetracycline resistance, <i>aphA-3</i>  =  kanamycin resistance, <i>ermC</i>  =  erythromycin resistance.</p><p>Plasmids used in this study.</p

Lipid A substitution profiles of meningococcal oxidoreductase mutants.

<p>Lipid A profiles of LOS extracted from <i>N. meningitidis</i> strain NMB (Panel A), NMBΔ<i>lptA::aadA</i> (Panel B), NMBΔ<i>NmdsbA1</i>/<i>NmdsbA2</i> (Panel C), NMBΔ<i>NmdsbA3</i> (Panel D) and NMBΔ<i>dsbA1/dsbA2/dsbA3</i> (Panel E) as determined by MALDI-TOF MS. <i>bis</i>-Phosphorylated hexaacylated lipid A (<i>m/z</i> = 1712), the <i>mono</i>-phosphorylated (<i>m/z</i> = 1632) and the <i>tri</i>-phosphorylated derivative (<i>m/z</i> = 1792) were detected in all strains. Strain NMB and the oxidoreductase mutants all expressed the mono-phosphorylated, <i>bis</i>-phosphorylated and <i>tri</i>-phosphorylated hexaacylated lipid A with a single PEA addition (<i>m/z</i> = 1755, <i>m/z</i> = 1835 and <i>m/z</i> = 1915). Consistent with the loss of LptA activity, NMBΔ<i>lptA::aadA</i> lacked these ions.</p

Bacterial strains and plasmids used in this study.

<p>*Nomenclature is derived from serological typing scheme for capsule polysaccharide (serogroup B):porin B variant (2B):porin A variant (P1.2,5):lipooligosaccharide immunotype (L2). Cassettes: <i>aadA</i>  =  spectinomycin resistance, <i>tetM</i>  =  tetracycline resistance, <i>aphA-3</i>  =  kanamycin resistance, <i>ermC</i>  =  erythromycin resistance.</p><p>+ ND =  not done.</p><p>Bacterial strains and plasmids used in this study.</p