Genomic Dissection of Bipolar Disorder and Schizophrenia, Including 28 Subphenotypes

publisher: Elsevier articletitle: Genomic Dissection of Bipolar Disorder and Schizophrenia, Including 28 Subphenotypes journaltitle: Cell articlelink: https://doi.org/10.1016/j.cell.2018.05.046 content_type: article copyright: © 2018 Elsevier Inc

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

A processed pseudogene contributes to apparent mule deer prion gene heterogeneity

Pathogenesis and transmission of the prion disorders (transmissible spongiform encephalopathies, TSEs) are mediated by a modified isoform of the prion protein (PrP). Prion protein gene (PRNP) alleles associated with relative susceptibility to TSE have been identified in sheep, humans and possibly elk. Comparable data have not been derived for mule deer, a species susceptible to the TSE chronic wasting disease (CWD). Initial analysis of the open reading frame (ORF) in exon 3 of the mule deer PRNP gene revealed polymorphisms in all 145 samples analyzed, with 10 potential polymorphic sites. Because 144/145 (99.3%) of the samples were heterozygous for a coding change (N/ S) at codon 138 (bp 412) and a non-coding polymorphism at bp 418, and individual deer with three or four different alleles were identified a possible gene duplication was indicated. Analysis of BAC clones containing mule deer PRNP genes revealed a full length functional gene and a processed pseudogene. The pseudogene was characteristic of previously described retroelements, in that it lacks introns and is flanked by repeat sequences. Three alleles of the functional gene were identified, with coding changes only at codons 20 (D/G) and 225 (S/F). Determination of PRNP functional gene alleles from 47 CWD-positive mule deer showed the predominant allele encoded 20D225S (frequency 0.85). When alleles were grouped by coding changes in the functional gene, four of the six possible peptide combinations were identified in infected deer. Three pseudogene alleles with coding changes in exon 3 were identified in the mule deer samples examined. Because the TSEs appear to be ‘‘protein only’’ disorders, the presence of an untranslated pseudogene is not expected to affect disease resistance. Therefore, selection of a genotyping method specific for the functional gene is critical for large-scale studies to identify the role of the PRNP gene in susceptibility to CWD in mule deer

Tick-borne transmission of two genetically distinct Anaplasma marginale strains following superinfection of the mammalian reservoir host

Strain superinfection affects the dynamics of epidemiological spread of pathogens through a host population. Superinfection has recently been shown to occur for two genetically distinct strains of the tick-borne pathogen Anaplasma marginale that encode distinctly different surface protein variants. Superinfected animals could serve as a reservoir for onward transmission of both strains if the tick vector is capable of acquiring and transmitting both strains. Whether competition among strains during development within the tick vector, which requires sequential invasion and replication events, limits colonization and subsequent transmission to a single strain is unknown. We tested this possibility by acquisition feeding Dermacentor andersoni ticks on a reservoir host superinfected with the genetically distinct St. Maries and EMPhi strains. Although the St. Maries strain consistently maintained higher bacteremia levels in the mammalian host and the EMPhi strain had an early advantage in colonization of the tick salivary glands, individual ticks were coinfected, and there was successful transmission of both strains. These results indicate that a genetically distinct A. marginale strain capable of superinfecting the mammalian host can subsequently be cotransmitted and become established within the host population despite the presence of an existing established strain

Identification of midgut and salivary glands as specific and distinct barriers to efficient tick-borne transmission of Anaplasma marginale

Understanding the determinants of efficient tick-borne microbial transmission is needed to better predict the emergence of highly transmissible pathogen strains and disease outbreaks. Although the basic developmental cycle of Anaplasma and Ehrlichia spp. within the tick has been delineated, there are marked differences in the ability of specific strains to be efficiently tick transmitted. Using the highly transmissible St. Maries strain of Anaplasma marginale in Dermacentor andersoni as a positive control and two unrelated nontransmissible strains, we identified distinct barriers to efficient transmission within the tick. The Mississippi strain was unable to establish infection at the level of the midgut epithelium despite successful ingestion of infected blood following acquisition feeding on a bacteremic animal host. This inability to colonize the midgut epithelium prevented subsequent development within the salivary glands and transmission. In contrast, A. marginale subsp. centrale colonized the midgut and then the salivary glands, replicating to a titer indistinguishable from that of the highly transmissible St. Maries strain and at least 100 times greater than that previously associated with successful transmission. Nonetheless, A. marginale subsp. centrale was not transmitted, even when a large number of infected ticks was used for transmission feeding. These results establish that there are at least two specific barriers to efficient tick-borne transmission, the midgut and salivary glands, and highlight the complexity of the pathogen-tick interaction

Transfected <i>Babesia bovis</i> Expressing a Tick GST as a Live Vector Vaccine

<div><p>The <i>Rhipicephalus microplus</i> tick is a notorious blood-feeding ectoparasite of livestock, especially cattle, responsible for massive losses in animal production. It is the main vector for transmission of pathogenic bacteria and parasites, including <i>Babesia bovis</i>, an intraerythrocytic apicomplexan protozoan parasite responsible for bovine Babesiosis. This study describes the development and testing of a live <i>B</i>. <i>bovis</i> vaccine expressing the protective tick antigen glutathione-S-transferase from <i>Haemaphysalis longicornis</i> (HlGST). The <i>B</i>. <i>bovis</i> S74-T3B parasites were electroporated with a plasmid containing the bidirectional <i>Ef-1α</i> (<i>elongation factor 1 alpha</i>) promoter of <i>B</i>. <i>bovis</i> controlling expression of two independent genes, the selectable marker <i>GFP-BSD</i> (<i>green fluorescent protein–blasticidin deaminase</i>), and <i>HlGST</i> fused to the <i>MSA-1</i> (<i>merozoite surface antigen 1</i>) signal peptide from <i>B</i>. <i>bovis</i>. Electroporation followed by blasticidin selection resulted in the emergence of a mixed <i>B</i>. <i>bovis</i> transfected line (termed HlGST) in <i>in vitro</i> cultures, containing parasites with distinct patterns of insertion of both exogenous genes, either in or outside the <i>Ef-1α</i> locus. A <i>B</i>. <i>bovis</i> clonal line termed HlGST-Cln expressing intracellular GFP and HlGST in the surface of merozoites was then derived from the mixed parasite line HlGST using a fluorescent activated cell sorter. Two independent calf immunization trials were performed via intravenous inoculation of the HlGST-Cln and a previously described control consisting of an irrelevant transfected clonal line of <i>B</i>. <i>bovis</i> designated GFP-Cln. The control GFP-Cln line contains a copy of the GFP-BSD gene inserted into the <i>Ef-1α</i> locus of <i>B</i>. <i>bovis</i> in an identical fashion as the HIGST-Cln parasites. All animals inoculated with the HlGST-Cln and GFP-Cln transfected parasites developed mild babesiosis. Tick egg fertility and fully engorged female tick weight was reduced significantly in <i>R</i>. <i>microplus</i> feeding on HlGST-Cln-immunized calves. Collectively, these data show the efficacy of a transfected HlGST-Cln <i>B</i>. <i>bovis</i> parasite to induce detectable anti-glutathione-S-transferase antibodies and a reduction in tick size and fecundity of <i>R</i>. <i>microplus</i> feeding in experimentally inoculated animals.</p></div

HlGST parasites immunofluorescence.

<p>Immunofluorescence assays using DAPI stained permeabilized or non-permeabilized free merozoites derived the from HIGST-Cln <i>B</i>. <i>bovis</i> cell line. Non-permeabilized free merozoites cells were incubated with anti-MSA-1 (Alexa Fluor 488) and anti-HlGST (Alexa Fluor 555). Non-permeabilized free merozoites were also incubated with anti-GFP (Alexa Fluor 488) and anti-HlGST (Alexa Fluor 555).Permeabilized merozoites were incubated with anti-GFP (Alexa Fluor 488) and anti-GST (Alexa Fluor 555), pre-immune rabbit serum (Alexa fluor 488), control anti-Tryp unrelated (Alexa Fluor 555). Columns represent DAPI, green (488nm), red (555nm) and green/red merged (488nm+555nm). The size bar is indicated on lower right image.</p

Characterization of transfected parasites.

<p>Two lines of transfected parasites HlGST1 and HlGST2 were generated by transfection of the T3B strain of <i>B</i>. <i>bovis</i> with plasmid <i>pMSASignal-HlGST-GFP-BSD</i> and analyzed in these experiments A) Comparison of the growth curves of non-transfected, control transfected (negative control electroporated with plasmid <i>pBS</i>, and unrelated positive control electroporated with plasmid <i>pEf-msa-1-Bm86ep-gfp-bsd</i>), and two lines of parasites electroporated with <i>pMSASignal-HlGST-GFP-BSD</i> (HlGST1 and HlGST2) after electroporation in the presence inhibitory doses of blasticidin. Blasticidin resistant parasites emerge ~16 days after the onset of selection only in the wells containing parasites electroporated with the <i>pMSASignal-HlGST-GFP-BSD</i> and <i>pEf-msa-1-Bm86ep-gfp-bsd</i> plasmids. B) Fluorescence microscopy of transfected parasites of the HlGST line (HlGST1 and 2, Upper panels), control GFP-<i>B</i>. <i>bovis</i> line (Unrelated) and non-transfected parasites (Lower panels).</p

Integration PCR analysis.

<p>Upper panel: Representation of the genome area including the transfected genes integrated into the genome of HlGST-Cln <i>B</i>. <i>bovis</i>. The localization of the regions hybridizing with the primers used in PCR is represented in the map by arrows. Primers were used for the amplification of EF-GST, HlGST, RAP-1, GFP-EF and GFP/BSD. Lower Panel: Agarose gel analysis of the PCR amplification products: Lane 1: HlGST 1 transfected <i>B</i>. <i>bovis</i> line; lane 2: HlGST 2 transfected <i>B</i>. <i>bovis</i>; lane 3: unrelated (GFP) transfected control <i>B</i>. <i>bovis</i>; lane 4: non-transfected <i>B</i>. <i>bovis</i>: lane 5: <i>MSASignal-HlGST-GFP-BSD</i> plasmid; lane 6: unrelated transfection control plasmid; lane 7: negative no DNA control.</p

Primers and MSA Signal Peptide template used in plasmid construction

<p>Primers and MSA Signal Peptide template used in plasmid construction</p