bluuebunny/crossref_metadata_2025_split huggingface dataset:
Recent Advances in Detection of Plasmodium ovale: Implications of Separation into the Two Species Plasmodium ovale wallikeri and Plasmodium ovale curtisi
Authors: Fuehrer Hans-Peter , Noedl Harald 
Published: October 2013

ABSTRACT

Recent molecular studies indicate that
Plasmodium ovale
malaria is caused by two closely related species of protozoan parasites, thereby imposing new challenges for detection and species differentiation. This minireview explores the potential value of innovative methods for the molecular diagnosis of malaria with a strong emphasis on the discrimination and genotyping of
P. ovale wallikeri
and
P. ovale curtisi
as well as tools for the simultaneous detection of
P. ovale
sp. An update for the widely used NP-1993 to NP-2005 (SSU rRNA) protocols for all human malaria parasites is discussed.
A Novel Subnucleocapsid Nanoplatform for Mucosal Vaccination against Influenza Virus That Targets the Ectodomain of Matrix Protein 2
Authors: Hervé Pierre-Louis , Raliou Mariam , Bourdieu Christiane , Dubuquoy Catherine , Petit-Camurdan Agnès , Bertho Nicolas , Eléouët Jean-François , Chevalier Christophe , Riffault Sabine 
Published: October 2013

ABSTRACT
In this study, subnucleocapsid nanorings formed by the recombinant nucleoprotein (N) of the respiratory syncytial virus were evaluated as a platform to anchor heterologous antigens. The ectodomain of the influenza virus A matrix protein 2 (M2e) is highly conserved and elicits protective antibodies when it is linked to an immunogenic carrier, making it a promising target to develop universal influenza vaccines. In this context, one or three M2e copies were genetically linked to the C terminus of N to produce N-M2e and N-3M2e chimeric recombinant nanorings. Mice were immunized intranasally with N-M2e or N-3M2e or with M2e or 3M2e control peptides. N-3M2e-vaccinated mice showed the strongest mucosal and systemic antibody responses. These mice presented a reduced viral load and minor weight loss, and all survived upon challenge with influenza virus A/PR8/34 (H1N1) (PR8). We compared the intranasal route to the subcutaneous route of N-3M2e immunization. Only the intranasal route induced a strong local IgA response and led to the protection of mice upon challenge. Finally, we demonstrated that the induction of anti-M2e antibodies by N-3M2e is not impaired by preexisting anti-N immunity. Overall, these results show that the N nanoring is a potent carrier for mucosal delivery of vaccinal antigens.
Adaptive Mutations in the Nuclear Export Protein of Human-Derived H5N1 Strains Facilitate a Polymerase Activity-Enhancing Conformation
Authors: Reuther Peter , Giese Sebastian , Götz Veronika , Kilb Normann , Mänz Benjamin , Brunotte Linda , Schwemmle Martin 
Published: October 2013

ABSTRACT

The nuclear export protein (NEP) (NS2) of the highly pathogenic human-derived H5N1 strain A/Thailand/1(KAN-1)/2004 with the adaptive mutation M16I greatly enhances the polymerase activity in human cells in a concentration-dependent manner. While low NEP levels enhance the polymerase activity, high levels are inhibitory. To gain insights into the underlying mechanism, we analyzed the effect of NEP deletion mutants on polymerase activity after reconstitution in human cells. This revealed that the polymerase-enhancing function of NEP resides in the C-terminal moiety and that removal of the last three amino acids completely abrogates this activity. Moreover, compared to full-length NEP, the C-terminal moiety alone exhibited significantly higher activity and seemed to be deregulated, since even the highest concentration did not result in an inhibition of polymerase activity. To determine transient interactions between the N- and C-terminal domains in
cis
, we fused both ends of NEP to a split click beetle luciferase and performed fragment complementation assays. With decreasing temperature, increased luciferase activity was observed, suggesting that intramolecular binding between the C- and N-terminal domains is preferentially stabilized at low temperatures. This stabilizing effect was significantly reduced with the adaptive mutation M16I or a combination of adaptive mutations (M16I, Y41C, and E75G), which further increased polymerase activity also at 34°C. We therefore propose a model in which the N-terminal moiety of NEP exerts an inhibitory function by back-folding to the C-terminal domain. In this model, adaptive mutations in NEP decrease binding between the C- and N-terminal domains, thereby allowing the protein to “open up” and become active already at a low temperature.
Activation of Influenza A Viruses by Host Proteases from Swine Airway Epithelium
Authors: Peitsch Catharina , Klenk Hans-Dieter , Garten Wolfgang , Böttcher-Friebertshäuser Eva 
Published: October 2013

ABSTRACT
Pigs are important natural hosts of influenza A viruses, and due to their susceptibility to swine, avian, and human viruses, they may serve as intermediate hosts supporting adaptation and genetic reassortment. Cleavage of the influenza virus surface glycoprotein hemagglutinin (HA) by host cell proteases is essential for viral infectivity. Most influenza viruses, including human and swine viruses, are activated at a monobasic HA cleavage site, and we previously identified TMPRSS2 and HAT to be relevant proteases present in human airways. We investigated the proteolytic activation of influenza viruses in primary porcine tracheal and bronchial epithelial cells (PTEC and PBEC, respectively). Human H1N1 and H3N2 viruses replicated efficiently in PTECs and PBECs, and viruses containing cleaved HA were released from infected cells. Moreover, the cells supported the proteolytic activation of HA at the stage of entry. We found that swine proteases homologous to TMPRSS2 and HAT, designated swTMPRSS2 and swAT, respectively, were expressed in several parts of the porcine respiratory tract. Both proteases cloned from primary PBECs were shown to activate HA with a monobasic cleavage site upon coexpression and support multicycle replication of influenza viruses. swAT was predominantly localized at the plasma membrane, where it was present as an active protease that mediated activation of incoming virus. In contrast, swTMPRSS2 accumulated in the trans-Golgi network, suggesting that it cleaves HA in this compartment. In conclusion, our data show that HA activation in porcine airways may occur by similar proteases and at similar stages of the viral life cycle as in human airways.
CD4 T Cell Help Is Limiting and Selective during the Primary B Cell Response to Influenza Virus Infection
Authors: Alam Shabnam , Knowlden Zackery A. G. , Sangster Mark Y. , Sant Andrea J. 
Published: October 2013

ABSTRACTInfluenza virus vaccination strategies are focused upon the elicitation of protective antibody responses through administration of viral protein through either inactivated virions or live attenuated virus. Often overlooked in this strategy is the CD4 T cell response: how it develops into memory, and how it may support future primary B cell responses to heterologous infection. Through the utilization of a peptide-priming regimen, this study describes a strategy for developing CD4 T cell memory with the capacity to robustly expand in the lung-draining lymph node after live influenza virus infection. Not only were frequencies of antigen-specific CD4 T cells enhanced, but these cells also supported an accelerated primary B cell response to influenza virus-derived protein, evidenced by high anti-nucleoprotein (NP) serum antibody titers early, while there is still active viral replication ongoing in the lung. NP-specific antibody-secreting cells and heightened frequencies of germinal center B cells and follicular T helper cells were also readily detectable in the draining lymph node. Surprisingly, a boosted memory CD4 T cell response was not sufficient to provide intermolecular help for antibody responses. Our study demonstrates that CD4 T cell help is selective and limiting to the primary antibody response to influenza virus infection and that preemptive priming of CD4 T cell help can promote effective and rapid conversion of naive B cells to mature antibody-secreting cells.
Crystal Structure of pb9, the Distal Tail Protein of Bacteriophage T5: a Conserved Structural Motif among All Siphophages
Authors: Flayhan Ali , Vellieux Frédéric M. D. , Lurz Rudi , Maury Olivier , Contreras-Martel Carlos , Girard Eric , Boulanger Pascale , Breyton Cécile 
Published: October 2013

ABSTRACT

The tail of
Caudovirales
bacteriophages serves as an adsorption device, a host cell wall-perforating machine, and a genome delivery pathway. In
Siphoviridae
, the assembly of the long and flexible tail is a highly cooperative and regulated process that is initiated from the proteins forming the distal tail tip complex. In Gram-positive-bacterium-infecting siphophages, the distal tail (Dit) protein has been structurally characterized and is proposed to represent a baseplate hub docking structure. It is organized as a hexameric ring that connects the tail tube and the adsorption device. In this study, we report the characterization of pb9, a tail tip protein of
Escherichia coli
bacteriophage T5. By immunolocalization, we show that pb9 is located in the upper part of the cone of the T5 tail tip, at the end of the tail tube. The crystal structure of pb9 reveals a two-domain protein. Domain A exhibits remarkable structural similarity with the N-terminal domain of known Dit proteins, while domain B adopts an oligosaccharide/oligonucleotide-binding fold (OB-fold) that is not shared by these proteins. We thus propose that pb9 is the Dit protein of T5, making it the first Dit protein described for a Gram-negative-bacterium-infecting siphophage. Multiple sequence alignments suggest that pb9 is a paradigm for a large family of Dit proteins of siphophages infecting mostly Gram-negative hosts. The modular structure of the Dit protein maintains the basic building block that would be conserved among all siphophages, combining it with a more divergent domain that might serve specific host adhesion properties.
The D10 Decapping Enzyme of Vaccinia Virus Contributes to Decay of Cellular and Viral mRNAs and to Virulence in Mice
Authors: Liu Shin-Wu , Wyatt Linda S. , Orandle Marlene S. , Minai Mahnaz , Moss Bernard 
Published: October 2013

ABSTRACT

Posttranscriptional mechanisms are important for regulation of cellular and viral gene expression. The presence of the 5′ cap structure m
7
G(5′)ppp(5′)Nm is a general feature of mRNAs that provides protection from exoribonuclease digestion and enhances translation. Vaccinia virus and other poxviruses encode enzymes for both cap synthesis and decapping. Decapping is mediated by two related enzymes, D9 and D10, which are synthesized before and after viral DNA replication, respectively. The timing of D10 synthesis correlates better with the shutdown of host gene expression, and deletion of this gene has been shown to cause persistence of host and viral mRNAs in infected cells. Here, we constructed specific mutant viruses in which translation of D10 was prevented by stop codons or activity of D10 was abrogated by catalytic site mutations, without other genomic alterations. Both mutants formed plaques of normal size and replicated to similar extents as the parental virus in monkey epithelial cells and mouse embryonic fibroblasts. The synthesis of viral proteins was slightly delayed, and cellular and viral mRNAs persisted longer in cells infected with the mutants compared to either the parental virus or clonal revertant. Despite the mild effects
in vitro
, both mutants were more attenuated than the revertants in intranasal and intraperitoneal mouse models, and less infectious virus was recovered from organs. In addition, there was less lung histopathology following intranasal infection with mutant viruses. These data suggest that the D10 decapping enzyme may help restrict antiviral responses by accelerating host mRNA degradation during poxvirus infection.
Discovery of Novel Ribonucleoside Analogs with Activity against Human Immunodeficiency Virus Type 1
Authors: Dapp Michael J. , Bonnac Laurent , Patterson Steven E. , Mansky Louis M. 
Published: October 2013

ABSTRACT

Reverse transcription is an important early step in retrovirus replication and is a key point targeted by evolutionarily conserved host restriction factors (e.g., APOBEC3G, SamHD1). Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) is a major target of antiretroviral drugs, and concerns regarding drug resistance and off-target effects have led to continued efforts for identifying novel approaches to targeting HIV-1 RT. Several observations, including those obtained from monocyte-derived macrophages, have argued that ribonucleotides and their analogs can, intriguingly, impact reverse transcription. For example, we have previously demonstrated that 5-azacytidine has its greatest antiviral potency during reverse transcription by enhancement of G-to-C transversion mutations. In the study described here, we investigated a panel of ribonucleoside analogs for their ability to affect HIV-1 replication during the reverse transcription process. We discovered five ribonucleosides—8-azaadenosine, formycin A, 3-deazauridine, 5-fluorocytidine, and 2′-
C
-methylcytidine—that possess anti-HIV-1 activity, and one of these (i.e., 3-deazauridine) has a primary antiviral mechanism that involves increased HIV-1 mutational loads, while quantitative PCR analysis determined that the others resulted in premature chain termination. Taken together, our findings provide the first demonstration of a series of ribonucleoside analogs that can target HIV-1 reverse transcription with primary antiretroviral mechanisms that include premature termination of viral DNA synthesis or enhanced viral mutagenesis.
Powerful Sequence Similarity Search Methods and In-Depth Manual Analyses Can Identify Remote Homologs in Many Apparently “Orphan” Viral Proteins
Authors: Kuchibhatla Durga B. , Sherman Westley A. , Chung Betty Y. W. , Cook Shelley , Schneider Georg , Eisenhaber Birgit , Karlin David G. 
Published: October 2013

ABSTRACT

The genome sequences of new viruses often contain many “orphan” or “taxon-specific” proteins apparently lacking homologs. However, because viral proteins evolve very fast, commonly used sequence similarity detection methods such as BLAST may overlook homologs. We analyzed a data set of proteins from RNA viruses characterized as “genus specific” by BLAST. More powerful methods developed recently, such as HHblits or HHpred (available through web-based, user-friendly interfaces), could detect distant homologs of a quarter of these proteins, suggesting that these methods should be used to annotate viral genomes. In-depth manual analyses of a subset of the remaining sequences, guided by contextual information such as taxonomy, gene order, or domain cooccurrence, identified distant homologs of another third. Thus, a combination of powerful automated methods and manual analyses can uncover distant homologs of many proteins thought to be orphans. We expect these methodological results to be also applicable to cellular organisms, since they generally evolve much more slowly than RNA viruses. As an application, we reanalyzed the genome of a bee pathogen,
Chronic bee paralysis virus
(CBPV). We could identify homologs of most of its proteins thought to be orphans; in each case, identifying homologs provided functional clues. We discovered that CBPV encodes a domain homologous to the
Alphavirus
methyltransferase-guanylyltransferase; a putative membrane protein, SP24, with homologs in unrelated insect viruses and insect-transmitted plant viruses having different morphologies (cileviruses, higreviruses, blunerviruses, negeviruses); and a putative virion glycoprotein, ORF2, also found in negeviruses. SP24 and ORF2 are probably major structural components of the virions.
Switching a Replication-Defective Adenoviral Vector into a Replication-Competent, Oncolytic Adenovirus
Authors: Nakashima Hiroshi , Chiocca E. Antonio 
Published: October 2013

ABSTRACT

The adenovirus immediate early gene E1A initiates the program of viral gene transcription and reprograms multiple aspects of cell function and behavior. For adenoviral (Ad) vector-mediated gene transfer and therapy approaches, where replication-defective (RD) gene transfer is required, E1A has thus been the primary target for deletions. For oncolytic gene therapy for cancer, where replication-competent (RC) Ad viral gene expression is needed, E1A has been either mutated or placed under tumor-specific transcriptional control. A novel Ad vector that initially infected target tumor cells in an RD manner for transgene expression but that could be “switched” into an RC, oncolytic state when needed might represent an advance in vector technology. Here, we report that we designed such an Ad vector (proAdΔ24.GFP), where initial Ad replication is silenced by a green fluorescent protein (GFP) transgene that blocks cytomegalovirus (CMV)-mediated transcription of E1A. This vector functions as a bona fide E1A-deleted RD vector in infected tumor cells. However, because the silencing GFP transgene is flanked by FLP recombination target (FRT) sites, we show that it can be efficiently excised by Flp recombinase site-specific recombination, either when
Flp
is expressed constitutively in cells or when it is provided in
trans
by coinfection with a second RD herpes simplex virus (HSV) amplicon vector. This switches the RD Ad, proAdΔ24.GFP, into a fully RC, oncolytic Ad (rAdΔ24) that lyses tumor cells in culture and generates oncolytic progeny virions.
In vivo
, coinfection of established flank tumors with the RD proAdΔ24.GFP and the RD
Flp
-bearing HSV1 amplicon leads to generation of RC, oncolytic rAdΔ24. In an orthotopic human glioma xenograft tumor model, coinjection of the RD proAdΔ24.GFP and the RD
Flp
-bearing HSV1 amplicon also led to a significant increase in animal survival, compared to controls. Therefore,
Flp
-FRT site-specific recombination can be applied to switch RD Ad into fully oncolytic RC Ad for tumor therapy and is potentially applicable to a variety of gene therapy approaches.
Kinase Control of Latent HIV-1 Infection: PIM-1 Kinase as a Major Contributor to HIV-1 Reactivation
Authors: Duverger Alexandra , Wolschendorf Frank , Anderson Joshua C. , Wagner Frederic , Bosque Alberto , Shishido Takao , Jones Jennifer , Planelles Vicente , Willey Christopher , Cron Randall Q. , Kutsch Olaf 
Published: October 2013

ABSTRACTDespite the clinical relevance of latent HIV-1 infection as a block to HIV-1 eradication, the molecular biology of HIV-1 latency remains incompletely understood. We recently demonstrated the presence of a gatekeeper kinase function that controls latent HIV-1 infection. Using kinase array analysis, we here expand on this finding and demonstrate that the kinase activity profile of latently HIV-1-infected T cells is altered relative to that of uninfected T cells. A ranking of altered kinases generated from these kinome profile data predicted PIM-1 kinase as a key switch involved in HIV-1 latency control. Using genetic and pharmacologic perturbation strategies, we demonstrate that PIM-1 activity is indeed required for HIV-1 reactivation in T cell lines and primary CD4 T cells. The presented results thus confirm that kinases are key contributors to HIV-1 latency control. In addition, through mutational studies we link the inhibitory effect of PIM-1 inhibitor IV (PIMi IV) on HIV-1 reactivation to an AP-1 motif in the CD28-responsive element of the HIV-1 long terminal repeat (LTR). The results expand our conceptual understanding of the dynamic interactions of the host cell and the latent HIV-1 integration event and position kinome profiling as a research tool to reveal novel molecular mechanisms that can eventually be targeted to therapeutically trigger HIV-1 reactivation.
Latent Gammaherpesvirus 68 Infection Induces Distinct Transcriptional Changes in Different Organs
Authors: Canny Susan P. , Goel Gautam , Reese Tiffany A. , Zhang Xin , Xavier Ramnik , Virgin Herbert W. 
Published: October 2013

ABSTRACT
Previous studies identified a role for latent herpesvirus infection in cross-protection against infection and exacerbation of chronic inflammatory diseases. Here, we identified more than 500 genes differentially expressed in spleens, livers, or brains of mice latently infected with gammaherpesvirus 68 and found that distinct sets of genes linked to different pathways were altered in the spleen compared to those in the liver. Several of the most differentially expressed latency-specific genes (e.g., the gamma interferon [IFN-γ], Cxcl9, and Ccl5 genes) are associated with known latency-specific phenotypes. Chronic herpesvirus infection, therefore, significantly alters the transcriptional status of host organs. We speculate that such changes may influence host physiology, the status of the immune system, and disease susceptibility.
Human Cytomegalovirus Infection of Langerhans-Type Dendritic Cells Does Not Require the Presence of the gH/gL/UL128-131A Complex and Is Blocked after Nuclear Deposition of Viral Genomes in Immature Cells
Authors: Lauron Elvin J. , Yu Dong , Fehr Anthony R. , Hertel Laura 
Published: October 2013

ABSTRACTHuman cytomegalovirus (CMV) enters its host via the oral and genital mucosae. Langerhans-type dendritic cells (LC) are the most abundant innate immune cells at these sites, where they constitute a first line of defense against a variety of pathogens. We previously showed that immature LC (iLC) are remarkably resistant to CMV infection, while mature LC (mLC) are more permissive, particularly when exposed to clinical-strain-like strains of CMV, which display a pentameric complex consisting of the viral glycoproteins gH, gL, UL128, UL130, and UL131A on their envelope. This complex was recently shown to be required for the infection of immature monocyte-derived dendritic cells. We thus sought to establish if the presence of this complex is also necessary for virion penetration of LC and if defects in entry might be the source of iLC resistance to CMV. Here we report that the efficiency of LC infection is reduced, but not completely abolished, in the absence of the pentameric complex. While virion penetration and nuclear deposition of viral genomes are not impaired in iLC, the transcription of the viral immediate early genes UL122 and UL123 and of the delayed early gene UL50 is substantially lower than that in mLC. Together, these data show that the UL128, UL130, and UL131A proteins are dispensable for CMV entry into LC and that progression of the viral cycle in iLC is restricted at the step of viral gene expression.