bluuebunny/crossref_metadata_2025 huggingface dataset:
Isoagglutinin reduction by a dedicated immunoaffinity chromatography step in the manufacturing process of human immunoglobulin products
Authors: Hoefferer Liane , Glauser Isabelle , Gaida Annette , Willimann Katharina , Marques Antunes Adriano , Siani Brigitte , Wymann Sandra , Widmer Eleonora , El Menyawi Ibrahim , Bolli Reinhard , Spycher Martin , Imboden Martin
Published: July 2015

BACKGROUND

The passive transfer of antibodies specific to blood groups A and B (also called isoagglutinins) contained in immunoglobulin (Ig)G products for intravenous administration (IVIG) is believed to be largely responsible for rare but sometimes serious IVIG‐related hemolytic events. We present in this work a modification of the manufacturing process of Privigen—a 10% l‐proline–stabilized IVIG product—that allows extensive reduction of isoagglutinin concentrations in the final product.

STUDY DESIGN AND METHODS

An additional immunoaffinity chromatography (IAC) step was introduced toward the end of the manufacturing process of Privigen. Isoagglutinin titers were measured using the indirect agglutination method and a published flow cytometry–based binding assay. Quality attributes, such as microorganism counts and concentration of endotoxins, IgG, IgA, IgM, aggregates, and so forth were measured using standardized procedures.

RESULTS

The introduction of an IAC step in the manufacturing process of Privigen resulted in an 88% to 90% reduction in isoagglutinins between the feed of the chromatography column and the flow‐through fraction. All other product quality attributes measured were nearly identical before and after IAC. This process modification resulted in a three‐titer‐step reduction in isoagglutinin levels in the final IgG product compared to Privigen lots produced by the unmodified process.

CONCLUSION

Introducing an isoagglutinin‐specific IAC step in the manufacturing process of Privigen is an efficient strategy for reduction of anti‐A and anti‐B titers. Such reductions might help minimize the risk of hemolytic events in patients receiving IVIG therapy.
Donor screening reduces the isoagglutinin titer in immunoglobulin products
Authors: Siani Brigitte , Willimann Katharina , Wymann Sandra , Marques Antunes Adriano , Widmer Eleonora
Published: July 2015

BACKGROUND

Hemolysis reaction is a rare class effect of therapy with intravenously administered human normal immunoglobulin (IVIG). Anti‐A/B isoagglutinins (isohemagglutinins) originating from donor plasma are considered a probable risk factor for hemolysis. We hypothesized that screening and exclusion of plasma obtained from donors with high isoagglutinin titers from the manufacturing process would produce a meaningful reduction of anti‐A/B isoagglutinin titers of the final IVIG product.

STUDY DESIGN AND METHODS

A donor screening method for anti‐A isoagglutinins using an automated indirect agglutination test (IAT) in gel cards was developed. Industry‐scale donor plasma pools and final IVIG product were prepared according to the manufacturing process of Privigen (human 10% liquid IVIG). Anti‐A/B isoagglutinin levels were measured by IAT, direct agglutination test, and a flow cytometry–based assay.

RESULTS

Screening of plasma from 705 randomly selected donors identified 6.8% donors with high anti‐A isoagglutinin titers in plasma. Exclusion of plasma from these donors resulted in a one‐titer‐step reduction of anti‐A isoagglutinin in laboratory‐scale pooled plasma. The same donor screening method applied to industry‐scale production resulted in exclusion of 5.1% of donors and produced a one‐titer‐step reduction of both anti‐A and anti‐B isoagglutinin titers in the final IVIG product.

CONCLUSION

Anti‐A/B isoagglutinin titers in IVIG products can be reduced on an industrial scale through implementation of anti‐A donor screening, which may lower the risk of hemolysis after IVIG therapy.
The role of inflammation in intravenous immune globulin–mediated hemolysis
Authors: Pendergrast Jacob , Willie‐Ramharack Kezia , Sampson Lorna , Laroche Vincent , Branch Donald R.
Published: July 2015

Intravenous immune globulin (IVIG) therapy has shown great success in a number of autoimmune and inflammatory conditions and its use continues to increase worldwide. There is growing awareness of significant side effects of high‐dose IVIG: however, particularly severe hemolysis in patients that are non–group O. It has been proposed that IVIG‐associated hemolysis may be heralded by an existing inflammatory condition. In the work presented herein, we have provided a review of the pathophysiology of inflammation, particularly as it applies in immune‐mediated red blood cell hemolysis, and a summary of previous publications that suggest an association between IVIG‐mediated hemolysis and a state of existing inflammation. In addition, preliminary results from a prospective study to address the mechanism of IVIG‐associated hemolysis are provided. These preliminary data support the idea of an existing inflammatory condition preceding overt hemolysis after high‐dose IVIG therapy that: 1) is restricted to non–group O patients, 2) is seen when using IVIG doses of more than 2 g/kg, 3) involves an activated mononuclear phagocyte system, 4) may be presaged by a significant increase in the anti‐inflammatory cytokine interleukin‐1 receptor agonist, and 5) is independent of secretor status.
The first case of foetal neonatal alloimmune thrombocytopenia caused by anti‐HPA‐1a antibodies in Asian population
Authors: Armawai M. I. , Yahya N. M. , Hassan A. , Matsuhashi M. , Santoso S. , Tsuno N. H.
Published: July 2015

Background and Objectives

The foetal neonatal platelet alloimmune thrombocytopenia (FNAIT) is one of the most frequent clinical implications of immunization against human platelet antigen (HPA). Maternal IgG platelet alloantibodies induce the destruction of foetal platelets leading to thrombocytopenia. In Caucasian, alloantibodies against HPA‐1a are responsible for the majority of severe FNAIT. As the frequency of HPA‐1bb homozygous individuals is very low in Asia, the clinical relevance of anti‐HPA‐1a alloantibodies is doubtful in this population.

Study design and methods

Recently, we studied retrospectively sera from mothers with suspected FNAIT in Malaysia, a multi‐ethnic country. Surprisingly, we found in one maternal serum antibodies which may react with HPA‐1a. In this study, this FNAIT case, suspected to be caused by the anti‐HPA‐1a alloantibody, was investigated in more details.

Results

Antibody screening analysis using HPA‐phenotyped platelets by the antigen capture assay, MAIPA, showed specific reaction with the platelets GPIIb/IIIa carrying HPA‐1a antigenic determinants. In the control experiment, this antibody did not react with GPIIb/IIIa from HPA‐1bb individuals. In addition, no reaction was observed with other HPAs expressed on GPIIb/IIIa, GPIb/IX, GPIa/IIa, CD 109 and HLA Class I, both by screening tests and by cross‐match analysis (maternal serum against paternal platelets), in MAIPA. This result could be supported by the genotyping analysis of the family members (mother HPA‐1bb, father and the index child HPA‐1a positive).

Conclusions

Although considered rare, this case shows for the first time the clinical relevance of anti‐HPA‐1a antibody in the pathogenesis of FNAIT among Asian population.
PREVALENCE AND SIGNIFICANCE OF AN ULTRASONOGRAPHIC COLONIC MUSCULARIS HYPERECHOIC BAND PARALLELING THE SEROSAL LAYER IN DOGS
Authors: Heng Hock Gan , Lim Chee Kin , Miller Margaret A. , Broman Meaghan M.
Published: July 2015

The muscularis layer of the canine colon has been reported to appear homogeneously hypoechoic on ultrasonography. A hyperechoic band in the muscularis layer paralleling the serosal surface has been observed by authors in routine canine abdominal ultrasound examinations. The purpose of this prospective and retrospective cross‐sectional study was to determine the prevalence of this lesion, characterize its ultrasonographic and postmortem histologic features, and correlate its presence with clinical signs of gastrointestinal disease. In the prospective study, all dogs that underwent routine abdominal ultrasonography by one of two observers during a 4‐week period were included without any exclusion criteria. One observer reviewed ultrasound images and recorded the presence or absence of this lesion and its distribution, e.g. focal (< 2 cm long) or diffuse (> 2 cm long). In the retrospective study, all dogs that had both abdominal ultrasonography and necropsy from January 2011 to December 2013 were included without any exclusion criteria. Histologic examinations were performed by two observers and Masson's trichrome stain was used to identify fibrous collagen. Prevalence for the hyperechoic band was 32% in the prospective and 4.8% in the retrospective sample populations, respectively. The hyperechoic band appeared as diffuse, focal, or a combination of both. Histologic sections were available for six dogs. In a few cases, the lesion corresponded to the presence of fibrous tissue in the myenteric plexus or in the tunica muscularis. None of the dogs had a history of diarrhea. Findings supported the hypothesis that a colonic muscularis hyperechoic band paralleling the serosal layer in dogs could be a normal variant rather than a marker of disease.
Large-area SiC membrane produced by plasma enhanced chemical vapor deposition at relatively high temperature
Authors: Liu Yu , Xie Changqing
Published: July 2015

Advances in the growth of silicon carbide (SiC) thin films with outstanding thermal and mechanical properties have received considerable attention. However, the fabrication of large-area free-standing SiC membrane still remains a challenge. Here, the authors report a plasma enhanced chemical vapor deposition process at a relatively high temperature to improve the free-standing SiC membrane area. A systematic study on the microstructural, mechanical, and optical properties of hydrogenated polycrystalline silicon carbide (poly-SiCx:H) thin films deposited at 600 °C with different annealing temperatures has been performed. In the as-deposited state, SiCx:H thin films show a polycrystalline structure. The crystallinity degree can be further improved with the increase of the postdeposition annealing temperature. The resulting process produced free-standing 2-μm-thick SiC membranes up to 70 mm in diameter with root mean square roughness of 3.384 nm and optical transparency of about 70% at 632.8 nm wavelength. The large-area SiC membranes made out of poly-SiCx:H thin films deposited at a relatively high temperature can be beneficial for a wide variety of applications, such as x-ray diffractive optical elements, optical and mechanical filtering, lithography mask, lightweight space telescopes, etc.
Spatial release from masking in children with bilateral cochlear implants and with normal hearing: Effect of target-interferer similarity
Authors: Misurelli Sara M. , Litovsky Ruth Y.
Published: July 2015

In complex auditory environments, it is often difficult to separate a target talker from interfering speech. For normal hearing (NH) adult listeners, similarity between the target and interfering speech leads to increased difficulty in separating them; that is, informational masking occurs due to confusability of the target and interferers. This study investigated performance of children with bilateral cochlear implants (BiCIs) when target and interferers were either same-sex (male) talkers, or different-sex talkers (male target, female interferer). Comparisons between children with BiCIs and NH, when matched for age, were also conducted. Speech intelligibility was measured for target and interferers spatially co-located, or spatially separated with the interferers positioned symmetrically (+90° and −90°) or asymmetrically (both at +90°, right). Spatial release from masking (SRM) was computed as the difference between co-located and separated conditions. Within group BiCI comparisons revealed that in the co-located condition speech intelligibility was worse with the same-sex vs different-sex stimuli. There was also a trend for more SRM with the same-sex vs different-sex stimuli. When comparing BiCI to NH listeners, SRM was larger for the NH groups, suggesting that NH children are better able to make use of spatial cues to improve speech understanding in noise.
Report prescribes strong medicine for WHO
Authors: Kupferschmidt Kai
Published: July 2015

Ebola failures show that difficult reforms are needed, an independent panel says.
Russia targets Western ties
Authors: Pokrovsky Vladimir
Published: July 2015

Crackdown on “foreign agents” and “undesirable” groups threatens private support for science.
The synthetic therapist
Authors: Bohannon John
Published: July 2015

Some people prefer to bare their souls to computers rather than to fellow humans.
Fears of an AI pioneer
Authors: Bohannon John
Published: July 2015

Stuart Russell argues that AI is as dangerous as nuclear weapons.
Metrics for land-scarce agriculture
Authors: DeFries Ruth , Fanzo Jessica , Remans Roseline , Palm Cheryl , Wood Stephen , Anderman Tal L.
Published: July 2015

Nutrient content must be better integrated into planning
Machine learning: Trends, perspectives, and prospects
Authors: Jordan M. I. , Mitchell T. M.
Published: July 2015

Machine learning addresses the question of how to build computers that improve automatically through experience. It is one of today’s most rapidly growing technical fields, lying at the intersection of computer science and statistics, and at the core of artificial intelligence and data science. Recent progress in machine learning has been driven both by the development of new learning algorithms and theory and by the ongoing explosion in the availability of online data and low-cost computation. The adoption of data-intensive machine-learning methods can be found throughout science, technology and commerce, leading to more evidence-based decision-making across many walks of life, including health care, manufacturing, education, financial modeling, policing, and marketing.
3D structure of individual nanocrystals in solution by electron microscopy
Authors: Park Jungwon , Elmlund Hans , Ercius Peter , Yuk Jong Min , Limmer David T. , Chen Qian , Kim Kwanpyo , Han Sang Hoon , Weitz David A. , Zettl A. , Alivisatos A. Paul
Published: July 2015

Looking at teeny tiny platinum particles

Electron microscopy is a powerful technique for taking snapshots of particles or images at near-atomic resolution. Park
et al.
studied free-floating platinum nanoparticles using electron microscopy and liquid cells (see the Perspective by Colliex). Using analytical techniques developed to study biological molecules, they reconstructed the threedimensional features of the Pt particles at near-atomic resolution. This approach has the scope to study a mixed population of particles one at a time and to study their synthesis as it occurs in solution.

Science
, this issue p.
290
; see also p.
232
Magnetic bubbles with a twist
Authors: Bergmann Kirsten von
Published: July 2015

Individual skyrmionic bubbles can be generated and moved at room temperature

[Also see Research Article by

Jiang
et al.

]
Worldwide evidence of a unimodal relationship between productivity and plant species richness
Authors: Fraser Lauchlan H. , Pither Jason , Jentsch Anke , Sternberg Marcelo , Zobel Martin , Askarizadeh Diana , Bartha Sandor , Beierkuhnlein Carl , Bennett Jonathan A. , Bittel Alex , Boldgiv Bazartseren , Boldrini Ilsi I. , Bork Edward , Brown Leslie , Cabido Marcelo , Cahill James , Carlyle Cameron N. , Campetella Giandiego , Chelli Stefano , Cohen Ofer , Csergo Anna-Maria , Díaz Sandra , Enrico Lucas , Ensing David , Fidelis Alessandra , Fridley Jason D. , Foster Bryan , Garris Heath , Goheen Jacob R. , Henry Hugh A. L. , Hohn Maria , Jouri Mohammad Hassan , Klironomos John , Koorem Kadri , Lawrence-Lodge Rachael , Long Ruijun , Manning Pete , Mitchell Randall , Moora Mari , Müller Sandra C. , Nabinger Carlos , Naseri Kamal , Overbeck Gerhard E. , Palmer Todd M. , Parsons Sheena , Pesek Mari , Pillar Valério D. , Pringle Robert M. , Roccaforte Kathy , Schmidt Amanda , Shang Zhanhuan , Stahlmann Reinhold , Stotz Gisela C. , Sugiyama Shu-ichi , Szentes Szilárd , Thompson Don , Tungalag Radnaakhand , Undrakhbold Sainbileg , van Rooyen Margaretha , Wellstein Camilla , Wilson J. Bastow , Zupo Talita
Published: July 2015

Grassland diversity and ecosystem productivity

The relationship between plant species diversity and ecosystem productivity is controversial. The debate concerns whether diversity peaks at intermediate levels of productivity—the so-called humped-back model—or whether there is no clear predictable relationship. Fraser
et al.
used a large, standardized, and geographically diverse sample of grasslands from six continents to confirm the validity and generality of the humped-back model. Their findings pave the way for a more mechanistic understanding of the factors controlling species diversity.

Science
, this issue p.
302
Computational rationality: A converging paradigm for intelligence in brains, minds, and machines
Authors: Gershman Samuel J. , Horvitz Eric J. , Tenenbaum Joshua B.
Published: July 2015

After growing up together, and mostly growing apart in the second half of the 20th century, the fields of artificial intelligence (AI), cognitive science, and neuroscience are reconverging on a shared view of the computational foundations of intelligence that promotes valuable cross-disciplinary exchanges on questions, methods, and results. We chart advances over the past several decades that address challenges of perception and action under uncertainty through the lens of computation. Advances include the development of representations and inferential procedures for large-scale probabilistic inference and machinery for enabling reflection and decisions about tradeoffs in effort, precision, and timeliness of computations. These tools are deployed toward the goal of computational rationality: identifying decisions with highest expected utility, while taking into consideration the costs of computation in complex real-world problems in which most relevant calculations can only be approximated. We highlight key concepts with examples that show the potential for interchange between computer science, cognitive science, and neuroscience.
Neutrophil-macrophage communication in inflammation and atherosclerosis
Authors: Nahrendorf Matthias , Swirski Filip K.
Published: July 2015

Neutrophils may license macrophages to respond to cholesterol crystals and drive inflammation that aggravates atherosclerosis

[Also see Report by

Warnatsch
et al.

]
Is biodiversity good for your health?
Authors: Keesing Felicia , Ostfeld Richard S.
Published: July 2015

Disease incidence is often lower in more diverse communities of plants and animals
Virtual love
Ex Machina
Alex Garland, director
DNA Films, 2015. 108 minutes.
Authors: Picard Rosalind W.
Published: July 2015

A young programmer falls for a humanoid robot, but is the feeling mutual?
Evaluation of Cross-Protection of a Lineage 1 West Nile Virus Inactivated Vaccine against Natural Infections from a Virulent Lineage 2 Strain in Horses, under Field Conditions
Authors: Chaintoutis Serafeim C. , Diakakis Nikolaos , Papanastassopoulou Maria , Banos Georgios , Dovas Chrysostomos I.
Published: July 2015

Although experimental data regarding cross-protection of horse West Nile virus (WNV) vaccines against lineage 2 infections exist, the cross-protective efficacy of these vaccines under field conditions has not been demonstrated. This study was conducted to evaluate the capability of an inactivated lineage 1 vaccine (Equip WNV) to protect against natural infections from the Nea Santa-Greece-2010 lineage 2 strain. In total, 185 WNV-seronegative horses in Thessaloniki, Greece, were selected during 2 consecutive years (2011 and 2012); 140 were immunized, and 45 were used as controls. Horses were examined for signs compatible with WNV infection. Neutralizing antibody titers against the Greek strain and the PaAn001/France lineage 1 strain were determined in immunized horses. WNV circulation was detected during both years in the study area. It was estimated that 37% and 27% of the horses were infected during 2011 and 2012, respectively. Three control animals developed clinical signs, and the WNV diagnosis was confirmed. Signs related to WNV infection were not observed in the vaccinated animals. The nonvaccinated animals had a 7.58% ± 1.82% higher chance of exhibiting signs than immunized animals (
P
< 0.05). Neutralizing antibodies raised against both strains in all immunized horses were detectable 1 month after the initial vaccination course. The cross-protective capacity of the lowest titer (1:40) was evident in 19 animals which were subsequently infected and did not exhibit signs. Neutralizing antibodies were detectable until the annual booster, when strong anamnestic responses were observed (geometrical mean titer ratio [GMTR] for lineage 1 of 30.2; GMTR for lineage 2 of 27.5). The results indicate that Equip WNV is capable of inducing cross-protection against natural infections from a virulent lineage 2 WNV strain in horses.
Genomic Epidemiology of a Protracted Hospital Outbreak Caused by a Toxin A-Negative Clostridium difficile Sublineage PCR Ribotype 017 Strain in London, England
Authors: Cairns M. D. , Preston M. D. , Lawley T. D. , Clark T. G. , Stabler R. A. , Wren B. W.
Published: July 2015

Clostridium difficile
remains the leading cause of nosocomial diarrhea worldwide, which is largely considered to be due to the production of two potent toxins: TcdA and TcdB. However, PCR ribotype (RT) 017, one of five clonal lineages of human virulent
C. difficile
, lacks TcdA expression but causes widespread disease. Whole-genome sequencing was applied to 35 isolates from hospitalized patients with
C. difficile
infection (CDI) and two environmental ward isolates in London, England. The phylogenetic analysis of single nucleotide polymorphisms (SNPs) revealed a clonal cluster of temporally variable isolates from a single hospital ward at University Hospital Lewisham (UHL) that were distinct from other London hospital isolates.
De novo
assembled genomes revealed a 49-kbp putative conjugative transposon exclusive to this hospital clonal cluster which would not be revealed by current typing methodologies. This study identified three sublineages of
C. difficile
RT017 that are circulating in London. Similar to the notorious RT027 lineage, which has caused global outbreaks of CDI since 2001, the lineage of toxin-defective RT017 strains appears to be continually evolving. By utilization of WGS technologies to identify SNPs and the evolution of clonal strains, the transmission of outbreaks caused by near-identical isolates can be retraced and identified.
Challenges in Preparation of Cumulative Antibiogram Reports for Community Hospitals
Authors: Moehring Rebekah W. , Hazen Kevin C. , Hawkins Myra R. , Drew Richard H. , Sexton Daniel J. , Anderson Deverick J.
Published: July 2015

Knowledge of local antimicrobial resistance is critical for management of infectious diseases. Community hospitals' compliance with Clinical and Laboratory Standards Institute (CLSI) guidance for creation of cumulative antibiograms is uncertain. This descriptive cohort study of antibiogram reporting practices included community hospitals enrolled in the Duke Infection Control Outreach Network. Cumulative antibiograms from 2012 were reviewed for criteria on reporting practices and compliance with CLSI guidelines. Microbiology personnel were sent a voluntary, electronic survey on antibiogram preparation practices. Data were compiled using descriptive statistics. Thirty-two of 37 (86%) hospitals provided antibiograms; 26 of 37 (70%) also provided survey responses. Twelve (38%) antibiograms specified methods used for compiling data and exclusion of duplicates. Eight (25%) reported only species with >30 isolates. Of the 24 that did not follow the 30-isolate rule, 3 (13%) included footnotes to indicate impaired statistical validity. Twenty (63%) reported at least 1 pathogen-drug combination not recommended for primary or supplemental testing per CLSI. Thirteen (41%) separately reported methicillin-resistant and -susceptible
Staphylococcus aureus
. Complete compliance with CLSI guidelines was observed in only 3 (9%) antibiograms. Survey respondents' self-assessment of full or partial compliance with CLSI guidelines was 50% and 15%, respectively; 33% reported uncertainty with CLSI guidelines. Full adherence to CLSI guidelines for hospital antibiograms was uncommon. Uncertainty about CLSI guidelines was common. Alternate strategies, such as regional antibiograms using pooled data and educational outreach efforts, are needed to provide reliable and appropriate susceptibility estimates for community hospitals.
Isothermal Detection of Mycoplasma pneumoniae Directly from Respiratory Clinical Specimens
Authors: Petrone Brianna L. , Wolff Bernard J. , DeLaney Alexandra A. , Diaz Maureen H. , Winchell Jonas M.
Published: July 2015

Mycoplasma pneumoniae
is a leading cause of community-acquired pneumonia (CAP) across patient populations of all ages. We have developed a loop-mediated isothermal amplification (LAMP) assay that enables rapid, low-cost detection of
M. pneumoniae
from nucleic acid extracts and directly from various respiratory specimen types. The assay implements calcein to facilitate simple visual readout of positive results in approximately 1 h, making it ideal for use in primary care facilities and resource-poor settings. The analytical sensitivity of the assay was determined to be 100 fg by testing serial dilutions of target DNA ranging from 1 ng to 1 fg per reaction, and no cross-reactivity was observed against 17 other
Mycoplasma
species, 27 common respiratory agents, or human DNA. We demonstrated the utility of this assay by testing nucleic acid extracts (
n
= 252) and unextracted respiratory specimens (
n
= 72) collected during
M. pneumoniae
outbreaks and sporadic cases occurring in the United States from February 2010 to January 2014. The sensitivity of the LAMP assay was 88.5% tested on extracted nucleic acid and 82.1% evaluated on unextracted clinical specimens compared to a validated real-time PCR test. Further optimization and improvements to this method may lead to the availability of a rapid, cost-efficient laboratory test for
M. pneumoniae
detection that is more widely available to primary care facilities, ultimately facilitating prompt detection and appropriate responses to potential
M. pneumoniae
outbreaks and clusters within the community.
Cladosporium Species Recovered from Clinical Samples in the United States
Authors: Sandoval-Denis Marcelo , Sutton Deanna A. , Martin-Vicente Adela , Cano-Lira José F. , Wiederhold Nathan , Guarro Josep , Gené Josepa
Published: July 2015

Cladosporiumspecies are ubiquitous, saprobic, dematiaceous fungi, only infrequently associated with human and animal opportunistic infections. We have studied a large set ofCladosporiumisolates recovered from clinical samples in the United States to ascertain the predominant species there in light of recent taxonomic changes in this genus and to determine whether some could possibly be rare potential pathogens. A total of 92 isolates were identified using phenotypic and molecular methods, which included sequence analysis of the internal transcribed spacer (ITS) region and a fragment of the large subunit (LSU) of the nuclear ribosomal DNA (rDNA), as well as fragments of the translation elongation factor 1 alpha (EF-1α) and actin (Act) genes. The most frequent species wasCladosporium halotolerans(14.8%), followed byC. tenuissimum(10.2%),C. subuliforme(5.7%), andC. pseudocladosporioides(4.5%). However, 39.8% of the isolates did not correspond to any known species and were deemed to comprise at least 17 new lineages forCladosporium. The most frequent anatomic site of isolation was the respiratory tract (54.5%), followed by superficial (28.4%) and deep tissues and fluids (14.7%). Species of the two recently described cladosporiumlike generaToxicocladosporiumandPenidiellaare reported for the first time from clinical samples.In vitrosusceptibility testing of 92 isolates against nine antifungal drugs showed a variety of results but high activity overall for the azoles, echinocandins, and terbinafine.
The Multiplicity of Cellular Infection Changes Depending on the Route of Cell Infection in a Plant Virus
Authors: Gutiérrez Serafín , Pirolles Elodie , Yvon Michel , Baecker Volker , Michalakis Yannis , Blanc Stéphane
Published: July 2015

The multiplicity of cellular infection (MOI) is the number of virus genomes of a given virus species that infect individual cells. This parameter chiefly impacts the severity of within-host population bottlenecks as well as the intensity of genetic exchange, competition, and complementation among viral genotypes. Only a few formal estimations of the MOI currently are available, and most theoretical reports have considered this parameter as constant within the infected host. Nevertheless, the colonization of a multicellular host is a complex process during which the MOI may dramatically change in different organs and at different stages of the infection. We have used both qualitative and quantitative approaches to analyze the MOI during the colonization of turnip plants by
Turnip mosaic virus
. Remarkably, different MOIs were observed at two phases of the systemic infection of a leaf. The MOI was very low in primary infections from virus circulating within the vasculature, generally leading to primary foci founded by a single genome. Each lineage then moved from cell to cell at a very high MOI. Despite this elevated MOI during cell-to-cell progression, coinfection of cells by lineages originating in different primary foci is severely limited by the rapid onset of a mechanism inhibiting secondary infection. Thus, our results unveil an intriguing colonization pattern where individual viral genomes initiate distinct lineages within a leaf. Kin genomes then massively coinfect cells, but coinfection by two distinct lineages is strictly limited.

IMPORTANCE
The MOI is the size of the viral population colonizing cells and defines major phenomena in virus evolution, like the intensity of genetic exchange and the size of within-host population bottlenecks. However, few studies have quantified the MOI, and most consider this parameter as constant during infection. Our results reveal that the MOI can depend largely on the route of cell infection in a systemically infected leaf. The MOI is usually one genome per cell when cells are infected from virus particles moving long distances in the vasculature, whereas it is much higher during subsequent cell-to-cell movement in mesophyll. However, a fast-acting superinfection exclusion prevents cell coinfection by merging populations originating from different primary foci within a leaf. This complex colonization pattern results in a situation where within-cell interactions are occurring almost exclusively among kin and explains the common but uncharacterized phenomenon of genotype spatial segregation in infected plants.
Replication of Tomato Yellow Leaf Curl Virus in Its Whitefly Vector, Bemisia tabaci
Authors: Pakkianathan Britto Cathrin , Kontsedalov Svetlana , Lebedev Galina , Mahadav Assaf , Zeidan Muhammad , Czosnek Henryk , Ghanim Murad
Published: July 2015

Tomato yellow leaf curl virus
(TYLCV) is a begomovirus transmitted exclusively by the whitefly
Bemisia tabaci
in a persistent, circulative manner. Replication of TYLCV in its vector remains controversial, and thus far, the virus has been considered to be nonpropagative. Following 8 h of acquisition on TYLCV-infected tomato plants or purified virions and then transfer to non-TYLCV-host cotton plants, the amounts of virus inside whitefly adults significantly increased (>2-fold) during the first few days and then continuously decreased, as measured by the amounts of genes on both virus DNA strands. Reported alterations in insect immune and defense responses upon virus retention led us to hypothesize a role for the immune response in suppressing virus replication. After virus acquisition, stress conditions were imposed on whiteflies, and the levels of three viral gene sequences were measured over time. When whiteflies were exposed to TYLCV and treatment with two different pesticides, the virus levels continuously increased. Upon exposure to heat stress, the virus levels gradually decreased, without any initial accumulation. Switching of whiteflies between pesticide, heat stress, and control treatments caused fluctuating increases and decreases in virus levels. Fluorescence
in situ
hybridization analysis confirmed these results and showed virus signals inside midgut epithelial cell nuclei. Combining the pesticide and heat treatments with virus acquisition had significant effects on fecundity. Altogether, our results demonstrate for the first time that a single-stranded DNA plant virus can replicate in its hemipteran vector.

IMPORTANCE
Plant viruses in agricultural crops are of great concern worldwide. Many of them are transmitted from infected to healthy plants by insects. Persistently transmitted viruses often have a complex association with their vectors; however, most are believed not to replicate within these vectors. Such replication is important, as it contributes to the virus's spread and can impact vector biology.
Tomato yellow leaf curl virus
(TYLCV) is a devastating begomovirus that infects tomatoes. It is persistently transmitted by the whitefly
Bemisia tabaci
but is believed not to replicate in the insect. To demonstrate that TYLCV is, in fact, propagative (i.e., it replicates in its insect host), we hypothesized that insect defenses play a role in suppressing virus replication. We thus exposed whitefly to pesticide and heat stress conditions to manipulate its physiology, and we showed that under such conditions, the virus is able to replicate and significantly influence the insect's fecundity.
2′-5′-Oligoadenylate Synthetase-Like Protein Inhibits Respiratory Syncytial Virus Replication and Is Targeted by the Viral Nonstructural Protein 1
Authors: Dhar Jayeeta , Cuevas Rolando A. , Goswami Ramansu , Zhu Jianzhong , Sarkar Saumendra N. , Barik Sailen
Published: July 2015

2′-5′-Oligoadenylate synthetase-like protein (OASL) is an interferon-inducible antiviral protein. Here we describe differential inhibitory activities of human OASL and the two mouse OASL homologs against respiratory syncytial virus (RSV) replication. Interestingly, nonstructural protein 1 (NS1) of RSV promoted proteasome-dependent degradation of specific OASL isoforms. We conclude that OASL acts as a cellular antiviral protein and that RSV NS1 suppresses this function to evade cellular innate immunity and allow virus growth.
Efficacy of the Herpes Simplex Virus 2 (HSV-2) Glycoprotein D/AS04 Vaccine against Genital HSV-2 and HSV-1 Infection and Disease in the Cotton Rat Sigmodon hispidus Model
Authors: Boukhvalova Marina , McKay Jamall , Mbaye Aissatou , Sanford-Crane Hannah , Blanco Jorge C. G. , Huber Ashley , Herold Betsy C.
Published: July 2015

Subunit vaccines based on the herpes simplex virus 2 (HSV-2) glycoprotein D (gD-2) have been the major focus of HSV-2 vaccine development for the past 2 decades. Based on the promising data generated in the guinea pig model, a formulation containing truncated gD-2, aluminum salt, and MPL (gD/AS04) advanced to clinical trials. The results of these trials, however, were unexpected, as the vaccine protected against HSV-1 infection but not against HSV-2. To address this discrepancy, we developed a Depot medroxyprogesterone acetate (DMPA)-treated cotton rat
Sigmodon hispidus
model of HSV-2 and HSV-1 genital infection. The severity of HSV-1 genital herpes was less than that of HSV-2 genital herpes in cotton rats, and yet the model allowed for comparative evaluation of gD/AS04 immunogenicity and efficacy. Cotton rats were intramuscularly vaccinated using a prime boost strategy with gD/AS04 (Simplirix vaccine) or control vaccine formulation (hepatitis B vaccine FENDrix) and subsequently challenged intravaginally with HSV-2 or HSV-1. The gD/AS04 vaccine was immunogenic in cotton rats and induced serum IgG directed against gD-2 and serum HSV-2 neutralizing antibodies but failed to efficiently protect against HSV-2 disease or to decrease the HSV-2 viral load. However, gD/AS04 significantly reduced vaginal titers of HSV-1 and better protected animals against HSV-1 compared to HSV-2 genital disease. The latter finding is generally consistent with the clinical outcome of the Herpevac trial of Simplirix. Passive transfer of serum from gD/AS04-immunized cotton rats conferred stronger protection against HSV-1 genital disease. These findings suggest the need for alternative vaccine strategies and the identification of new correlates of protection.

IMPORTANCE
In spite of the high health burden of genital herpes, there is still no effective intervention against the disease. The significant gap in knowledge on genital herpes pathogenesis has been further highlighted by the recent failure of GSK HSV-2 vaccine Simplirix (gD/AS04) to protect humans against HSV-2 and the surprising finding that the vaccine protected against HSV-1 genital herpes instead. In this study, we report that gD/AS04 has higher efficacy against HSV-1 compared to HSV-2 genital herpes in the novel DMPA-synchronized cotton rat model of HSV-1 and HSV-2 infection. The findings help explain the results of the Simplirix trial.
Distribution and Redistribution of HIV-1 Nucleocapsid Protein in Immature, Mature, and Integrase-Inhibited Virions: a Role for Integrase in Maturation
Authors: Fontana Juan , Jurado Kellie A. , Cheng Naiqian , Ly Ngoc L. , Fuchs James R. , Gorelick Robert J. , Engelman Alan N. , Steven Alasdair C.
Published: July 2015

During virion maturation, HIV-1 capsid protein assembles into a conical core containing the viral ribonucleoprotein (vRNP) complex, thought to be composed mainly of the viral RNA and nucleocapsid protein (NC). After infection, the viral RNA is reverse transcribed into double-stranded DNA, which is then incorporated into host chromosomes by integrase (IN) catalysis. Certain IN mutations (class II) and antiviral drugs (allosteric IN inhibitors [ALLINIs]) adversely affect maturation, resulting in virions that contain “eccentric condensates,” electron-dense aggregates located outside seemingly empty capsids. Here we demonstrate that in addition to this mislocalization of electron density, a class II IN mutation and ALLINIs each increase the fraction of virions with malformed capsids (from ∼12% to ∼53%). Eccentric condensates have a high NC content, as demonstrated by “tomo-bubblegram” imaging, a novel labeling technique that exploits the susceptibility of NC to radiation damage. Tomo-bubblegrams also localized NC inside wild-type cores and lining the spherical Gag shell in immature virions. We conclude that eccentric condensates represent nonpackaged vRNPs and that either genetic or pharmacological inhibition of IN can impair vRNP incorporation into mature cores. Supplying IN in
trans
as part of a Vpr-IN fusion protein partially restored the formation of conical cores with internal electron density and the infectivity of a class II IN deletion mutant virus. Moreover, the ability of ALLINIs to induce eccentric condensate formation required both IN and viral RNA. Based on these observations, we propose a role for IN in initiating core morphogenesis and vRNP incorporation into the mature core during HIV-1 maturation.

IMPORTANCE
Maturation, a process essential for HIV-1 infectivity, involves core assembly, whereby the viral ribonucleoprotein (vRNP, composed of vRNA and nucleocapsid protein [NC]) is packaged into a conical capsid. Allosteric integrase inhibitors (ALLINIs) affect multiple viral processes. We have characterized ALLINIs and integrase mutants that have the same phenotype. First, by comparing the effects of ALLINIs on several steps of the viral cycle, we show that inhibition of maturation accounts for compound potency. Second, by using cryoelectron tomography, we find that ALLINIs impair conical capsid assembly. Third, by developing tomo-bubblegram imaging, which specifically labels NC protein, we find that ALLINIs block vRNP packaging; instead, vRNPs form “eccentric condensates” outside the core. Fourth, malformed cores, typical of integrase-deleted virus, are partially replaced by conical cores when integrase is supplied in
trans
. Fifth, vRNA is necessary for ALLINI-induced eccentric condensate formation. These observations suggest that integrase is involved in capsid morphogenesis and vRNP packaging.
HIV Vpu Interferes with NF-κB Activity but Not with Interferon Regulatory Factor 3
Authors: Manganaro Lara , de Castro Elisa , Maestre Ana M. , Olivieri Kevin , García-Sastre Adolfo , Fernandez-Sesma Ana , Simon Viviana
Published: July 2015

The accessory HIV protein Vpu inhibits a number of cellular pathways that trigger host innate restriction mechanisms. HIV Vpu-mediated degradation of tetherin allows efficient particle release and hampers the activation of the NF-κB pathway thereby limiting the expression of proinflammatory genes. In addition, Vpu reduces cell surface expression of several cellular molecules such as newly synthesized CD4. However, the role of HIV Vpu in regulating the type 1 interferon response to viral infection by degradation of the interferon regulatory factor 3 (IRF3) has been subject of conflicting reports. We therefore systematically investigated the expression of IRF3 in primary CD4
+
T cells and macrophages infected with HIV at different time points. In addition, we also tested the ability of Vpu to interfere with innate immune signaling pathways such as the NF-κB and the IRF3 pathways. We report here that HIV Vpu failed to degrade IRF3 in infected primary cells. Moreover, we observed that HIV NL4.3 Vpu had no effect on IRF3-dependent gene expression in reporter assays. On the other hand, HIV NL4.3 Vpu downmodulated NF-κB-dependent transcription. Mutation of two serines (positions 52 and 56) involved in the binding of NL4.3 Vpu to the βTrCP ubiquitin ligase abolishes its ability to inhibit NF-κB activity. Taken together, these results suggest that HIV Vpu regulates antiviral innate response in primary human cells by acting specifically on the NF-κB pathway.

IMPORTANCE
HIV Vpu plays a pivotal role in enhancing HIV infection by counteraction of Tetherin. However, Vpu also regulates host response to HIV infection by hampering the type 1 interferon response. The molecular mechanism by which Vpu inhibits the interferon response is still controversial. Here we report that Vpu affects interferon expression by inhibiting NF-κB activity without affecting IRF3 levels or activity. These data suggest that Vpu facilitates HIV infection by regulating NF-κB transcription to levels sufficient for viral transcription while limiting cellular responses to infection.
Acylation of Biomolecules in Prokaryotes: a Widespread Strategy for the Control of Biological Function and Metabolic Stress
Authors: Hentchel Kristy L. , Escalante-Semerena Jorge C.
Published: July 2015

Acylation of biomolecules (e.g., proteins and small molecules) is a process that occurs in cells of all domains of life and has emerged as a critical mechanism for the control of many aspects of cellular physiology, including chromatin maintenance, transcriptional regulation, primary metabolism, cell structure, and likely other cellular processes. Although this review focuses on the use of acetyl moieties to modify a protein or small molecule, it is clear that cells can use many weak organic acids (e.g., short-, medium-, and long-chain mono- and dicarboxylic aliphatics and aromatics) to modify a large suite of targets. Acetylation of biomolecules has been studied for decades within the context of histone-dependent regulation of gene expression and antibiotic resistance. It was not until the early 2000s that the connection between metabolism, physiology, and protein acetylation was reported. This was the first instance of a metabolic enzyme (acetyl coenzyme A [acetyl-CoA] synthetase) whose activity was controlled by acetylation via a regulatory system responsive to physiological cues. The above-mentioned system was comprised of an acyltransferase and a partner deacylase. Given the reversibility of the acylation process, this system is also referred to as
r
eversible
l
ysine
a
cylation (RLA). A wealth of information has been obtained since the discovery of RLA in prokaryotes, and we are just beginning to visualize the extent of the impact that this regulatory system has on cell function.
Birefringence of Collagen Fibres in Rat Calcaneal Tendons Treated with Acupuncture during Three Phases of Healing
Authors: Almeida Marcos dos Santos , Oliveira Letícia Prado , Vieira Cristiano Pedrozo , Guerra Flávia Da Ré , Pimentel Edson Rosa
Published: July 2015

Background

Birefringence is an optical anisotropy that is investigated by polarisation microscopy, and has been valuable for the study of the oriented organisation of collagen fibres in tendons. However, the application of this technology to evaluate the effect of different acupuncture points during tendon healing has not yet been described.

Objectives

To evaluate the concentration of non-collagenous proteins (NCP) and birefringence in rat calcaneal tendons following injury during the three different phases of healing: inflammatory (7th day), proliferative (14th day), and remodelling (21st day).

Methods

Tendons of 120 Wistar rats were tenotomised and left untreated (teno group, n=24), treated with manual acupuncture at ST36 (ST36 group, n=24), BL57 (BL57 group, n=24) or ST36+BL57 (SB group, n=24), or treated with electroacupuncture at ST36+BL57 (EA group, n=24). Tendon samples were collected at 7, 14 and 21 days after injury (n=8 per group). NCP concentrations were measured using the Bradford method (n=4 each) and birefringence was examined using polarisation microscopy and image analysis (n=4 each). Comparison was also made with healthy (non-tenotomised) tendons in a subgroup of rats (n=4 each).

Results

Manual acupuncture at ST36 and BL57 increased molecular organisation of collagen fibres on day 14 and 21 after injury. Isolated use of BL57 and ST36 also increased collagen fibre organisation when examined on day 14 and 21, respectively. No significant increase in NCP concentration was observed in any of the treated tenotomised groups.

Conclusions

Acupuncture, through putative anti-inflammatory and mechanotransductor effects, may have a role in strengthening tendons and increasing resistance to re-rupture.
A trimodality approach in the management of metastatic low-grade epithelioid hemangioendothelioma of the bone
Authors: Saste Abhijit , Cabrera Fernandez Diego Felipe , Gulati Rohit , Gamalski Steven
Published: July 2015

A 29-year-old man presented with a 2-week history of severe pain in the left foot with no preceding history of trauma. A left foot radiograph demonstrated a cortical lucency in the mid-distal shaft of the third metatarsal bone. MRI of the left foot showed an expansile lesion in the same location. A staging bone scan showed a focal uptake in the known lucency in the left third metatarsal and in the proximal left femur. A subsequent left hip radiograph demonstrated a lucency in the intertrochanteric region. CT scan of the chest, abdomen and pelvis was unremarkable. A biopsy of the left third metatarsal expansile lesion performed during an incision and curettage procedure revealed an epithelioid haemangioma (EHE) of the bone. MRI of the left hip performed in response to the findings on the bone scan showed metastatic disease in the left intertrochanteric region. A prophylactic left hip fixation surgery with an interlocking intramedullary femoral nail was therefore undertaken to avoid a pathological fracture of the left hip from the metastatic disease. Simultaneously, a left hip biopsy was performed, which also revealed an EHE. The patient underwent external beam radiation to the left femoral head and neck. This was followed by fractionated radiosurgery to the left third metatarsal. Once the left foot wound had healed, the patient subsequently received four cycles of doxorubicin and ifosfamide. A restaging positron emission tomography CT carried out after completion of therapy showed no metabolic evidence of residual primary tumour or metastasis. More than 2 years after completing his trimodality therapy, the patient remains fully functional and symptom free.