title: Reference gene selection for normalization of RT-qPCR gene expression data from Actinidia deliciosa leaves infected with Pseudomonas syringae pv. actinidiae

text: Normalization of data, by choosing the appropriate reference genes (RGs), is fundamental for obtaining reliable results in reverse transcription-quantitative PCR (RT-qPCR). In this study, we assessed Actinidia deliciosa leaves inoculated with two doses of Pseudomonas syringae pv. actinidiae during a period of 13 days for the expression profile of nine candidate RGs. Their expression stability was calculated using four algorithms: geNorm, NormFinder, BestKeeper and the deltaCt method. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and protein phosphatase 2A (PP2A) were the most stable genes, while β-tubulin and 7s-globulin were the less stable. Expression analysis of three target genes, chosen for RGs validation, encoding the reactive oxygen species scavenging enzymes ascorbate peroxidase (APX), superoxide dismutase (SOD) and catalase (CAT) indicated that a combination of stable RGs, such as GAPDH and PP2A, can lead to an accurate quantification of the expression levels of such target genes. The APX level varied during the experiment time course and according to the inoculum doses, whereas both SOD and CAT resulted down-regulated during the first four days, and up-regulated afterwards, irrespective of inoculum dose. These results can be useful for better elucidating the molecular interaction in the A. deliciosa/P. s. pv. actinidiae pathosystem and for RGs selection in bacteria-plant pathosystems.


title: Hsp70 Isoforms Are Essential for the Formation of Kaposi’s Sarcoma-Associated Herpesvirus Replication and Transcription Compartments

text: Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic herpesvirus associated with various AIDS-related malignancies. Like other herpesviruses, multiple processes required for KSHV lytic replication, including viral transcription, viral DNA synthesis and capsid assembly occur in virus-induced intranuclear structures, termed replication and transcription compartments (RTCs). Here we utilised a novel methodology, combining subcellular fractionation and quantitative proteomics, to identify cellular proteins which are recruited to KSHV-induced RTCs and thus play a key role in KSHV lytic replication. We show that several isoforms of the HSP70 chaperone family, Hsc70 and iHsp70, are redistributed from the cytoplasm into the nucleus coinciding with the initial formation of KSHV-induced RTCs. We demonstrate that nuclear chaperone foci are dynamic, initially forming adjacent to newly formed KSHV RTCs, however during later time points the chaperones move within KSHV RTCs and completely co-localise with actively replicating viral DNA. The functional significance of Hsp70 isoforms recruitment into KSHV RTCs was also examined using the specific Hsp70 isoform small molecule inhibitor, VER-155008. Intriguingly, results highlight an essential role of Hsp70 isoforms in the KSHV replication cycle independent of protein stability and maturation. Notably, inhibition of Hsp70 isoforms precluded KSHV RTC formation and RNA polymerase II (RNAPII) relocalisation to the viral genome leading to the abolishment of global KSHV transcription and subsequent viral protein synthesis and DNA replication. These new findings have revealed novel mechanisms that regulate KSHV lytic replication and highlight the potential of HSP70 inhibitors as novel antiviral agents.


title: Rapid drop in the reproduction number during the Ebola outbreak in the Democratic Republic of Congo

text: The Democratic Republic of Congo (DRC) experienced a confined rural outbreak of Ebola virus disease (EVD) with 69 reported cases from July to October 2014. Understanding the transmission dynamics during the outbreak can provide important information for anticipating and controlling future EVD epidemics. I fitted an EVD transmission model to previously published data of this outbreak and estimated the basic reproduction number R(0) = 5.2 (95% CI [4.0–6.7]). The model suggests that the net reproduction number R(t) fell below unity 28 days (95% CI [25–34] days) after the onset of symptoms in the index case. This study adds to previous epidemiological descriptions of the 2014 EVD outbreak in DRC, and is consistent with the notion that a rapid implementation of control interventions helped reduce further spread.


title: Dynamics of ADAM17-Mediated Shedding of ACE2 Applied to Pancreatic Islets of Male db/db Mice

text: Angiotensin-converting enzyme 2 (ACE2) gene therapy aimed at counteracting pancreatic ACE2 depletion improves glucose regulation in two diabetic mouse models: db/db mice and angiotensin II-infused mice. A disintegrin and metalloproteinase 17 (ADAM17) can cause shedding of ACE2 from the cell membrane. The aim of our studies was to determine whether ADAM17 depletes ACE2 levels in pancreatic islets and β-cells. Dynamics of ADAM17-mediated ACE2 shedding were investigated in 832/13 insulinoma cells. Within a wide range of ACE2 expression levels, including the level observed in mouse pancreatic islets, overexpression of ADAM17 increases shed ACE2 and decreases cellular ACE2 levels. We provide a mathematical description of shed and cellular ACE2 activities as a function of the ADAM17 activity. The effect of ADAM17 on the cellular ACE2 content was relatively modest with an absolute control strength value less than 0.25 and approaching 0 at low ADAM17 activities. Although we found that ADAM17 and ACE2 are both expressed in pancreatic islets, the β-cell is not the major cell type expressing ACE2 in islets. During diabetes progression in 8-, 12-, and 15-week-old db/db mice, ACE2 mRNA and ACE2 activity levels in pancreatic islets were not decreased over time nor significantly decreased compared with nondiabetic db/m mice. Levels of ADAM17 mRNA and ADAM17 activity were also not significantly changed. Inhibiting basal ADAM17 activity in mouse islets failed to affect ACE2 levels. We conclude that whereas ADAM17 has the ability to shed ACE2, ADAM17 does not deplete ACE2 from pancreatic islets in diabetic db/db mice.


title: Chikungunya nsP2 protease is not a papain-like cysteine protease and the catalytic dyad cysteine is interchangeable with a proximal serine

text: Chikungunya virus is the pathogenic alphavirus that causes chikungunya fever in humans. In the last decade millions of cases have been reported around the world from Africa to Asia to the Americas. The alphavirus nsP2 protein is multifunctional and is considered to be pivotal to viral replication, as the nsP2 protease activity is critical for proteolytic processing of the viral polyprotein during replication. Classically the alphavirus nsP2 protease is thought to be papain-like with the enzyme reaction proceeding through a cysteine/histidine catalytic dyad. We performed structure-function studies on the chikungunya nsP2 protease and show that the enzyme is not papain-like. Characterization of the catalytic dyad cysteine residue enabled us to identify a nearby serine that is catalytically interchangeable with the dyad cysteine residue. The enzyme retains activity upon alanine replacement of either residue but a replacement of both cysteine and serine residues results in no detectable activity. Protein dynamics appears to allow the use of either the cysteine or the serine residue in catalysis. This switchable dyad residue has not been previously reported for alphavirus nsP2 proteases and would have a major impact on the nsP2 protease as an anti-viral target.


title: Sex-specific regulation of chemokine Cxcl5/6 controls neutrophil recruitment and tissue injury in acute inflammatory states

text: BACKGROUND: Tissue infiltration by neutrophils during acute inflammatory states causes substantial tissue injury. While the magnitude of tissue neutrophil accumulation in innate immune responses is profoundly greater in males than females, fundamental aspects of the molecular mechanisms underlying these sex differences remain largely unknown. METHODS: We investigated sex differences in neutrophil stimulation and recruitment in ischemia/reperfusion (I/R; mesenteric or renal) or carrageenan pleurisy in rats or mice, as well as skin injury in human volunteers. The induction of potent chemoattractive mediators (chemokines) and neutrophil adhesion molecules were measured by real-time PCR, flow cytometry, and protein assays. RESULTS: Mesenteric I/R in age-matched Wistar rats resulted in substantially more neutrophil accumulation and tissue injury at 2 h reperfusion in males than females. Using intravital microscopy, we show that the immediate (<30 min) neutrophil response to I/R is similar in males and females but that prolonged neutrophil recruitment occurs in males at sites local and distal to inflammatory insult partly due to an increase in circulating neutrophil populations with elevated surface expression of adhesion molecules. Sex differences in neutrophil kinetics were correlated with sustained induction of chemokine Cxcl5 in the tissue, circulation, and bone marrow of males but not females. Furthermore, blockade of Cxcl5 in males prior to ischemia resulted in neutrophil responses that were similar in magnitude to those in females. Conversely, administration of Cxcl5 to males in the absence of I/R was sufficient to increase levels of systemic neutrophils. Cxcl5 treatment of bone marrow neutrophils in vitro caused substantial induction of neutrophil-mobilizing cytokine granulocyte colony-stimulating factor (GCSF) and expression of β2 integrin that accounts for sexual dimorphism in circulating neutrophil populations in I/R. Moreover, male Cxcl5-stimulated bone marrow neutrophils had an increased capacity to adhere to β2 integrin ligand ICAM-1, implicating a greater sensitivity of male leukocytes to Cxcl5-mediated activation. Differential induction of Cxcl5 (human CXCL6) between the sexes was also evident in murine renal I/R, rat pleurisy, and human skin blisters and correlated with the magnitude of neutrophil accumulation in tissues. CONCLUSIONS: Our study reveals that sex-specific induction of chemokine Cxcl5/CXCL6 contributes to sexual dimorphism in neutrophil recruitment in diverse acute inflammatory responses partly due to increased stimulation and trafficking of bone marrow neutrophils in males. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13293-015-0047-5) contains supplementary material, which is available to authorized users.


title: M1 of Murine Gamma-Herpesvirus 68 Induces Endoplasmic Reticulum Chaperone Production

text: Viruses rely on host chaperone network to support their infection. In particular, the endoplasmic reticulum (ER) resident chaperones play key roles in synthesizing and processing viral proteins. Influx of a large amount of foreign proteins exhausts the folding capacity in ER and triggers the unfolded protein response (UPR). A fully-executed UPR comprises signaling pathways that induce ER folding chaperones, increase protein degradation, block new protein synthesis and may eventually activate apoptosis, presenting both opportunities and threats to the virus. Here, we define a role of the MHV-68M1 gene in differential modulation of UPR pathways to enhance ER chaperone production. Ectopic expression of M1 markedly induces ER chaperone genes and expansion of ER. The M1 protein accumulates in ER during infection and this localization is indispensable for its function, suggesting M1 acts from the ER. We found that M1 protein selectively induces the chaperon-producing pathways (IRE1, ATF6) while, interestingly, sparing the translation-blocking arm (PERK). We identified, for the first time, a viral factor capable of selectively intervening the initiation of ER stress signaling to induce chaperon production. This finding provides a unique opportunity of using viral protein as a tool to define the activation mechanisms of individual UPR pathways.


title: Immortalization of Fetal Bovine Colon Epithelial Cells by Expression of Human Cyclin D1, Mutant Cyclin Dependent Kinase 4, and Telomerase Reverse Transcriptase: An In Vitro Model for Bacterial Infection

text: Cattle are the economically important animals in human society. They are essential for the production of livestock products such as milk and meats. The production efficiency of livestock products is negatively impacted by infection with zoonotic pathogens. To prevent and control infectious diseases, it is important to understand the interaction between cattle tissue and pathogenic bacteria. In this study, we established an in vitro infection model of an immortalized bovine colon-derived epithelial cell line by transducing the cells with lentiviral vectors containing genes encoding cell cycle regulators cyclin D1, mutant cyclin dependent kinase 4 (CDK4), and human telomerase reverse transcriptase (TERT). The established cell line showed continuous cell proliferation, expression of epithelial markers, and an intact karyotype, indicating that the cells maintained their original nature as colon-derived epithelium. Furthermore, we exposed the established cell line to two strains of Salmonella enterica and EHEC. Interestingly, S. Typhimurium showed higher affinity for the established cell line and invaded the cytoplasm than S. Enteritidis. Quantitative RT-PCR revealed that gene expression of Toll-like receptor 1 (TLR1), TLR 2 and TLR 3, whereas TLR 4, 5 and 6 were not detectable in established cells. Our established immortalized colon-derived epithelial cell should be a useful tool for studies evaluating the molecular mechanisms underlying bacterial infection.


title: Proteomic Analysis of the Vitreous following Experimental Retinal Detachment in Rabbits

text: Purpose. The pathogenesis of rhegmatogenous retinal detachment (RRD) remains incompletely understood, with no clinically effective treatment for potentially severe complications such as photoreceptor cell death and proliferative vitreoretinopathy. Here we investigate the protein profile of the vitreous following experimental retinal detachment using a comparative proteomic based approach. Materials and Methods. Retinal detachment was created in the right eyes of six New Zealand red pigmented rabbits. Sham surgery was undertaken in five other rabbits that were used as controls. After seven days the eyes were enucleated and the vitreous was removed. The vitreous samples were evaluated with two-dimensional polyacrylamide gel electrophoresis and the differentially expressed proteins were identified with tandem mass spectrometry. Results. Ten protein spots were found to be at least twofold differentially expressed when comparing the vitreous samples of the sham and retinal detachment surgery groups. Protein spots that were upregulated in the vitreous following retinal detachment were identified as albumin fragments, and those downregulated were found to be peroxiredoxin 2, collagen-Iα1 fragment, and α-1-antiproteinase F. Conclusions. Proteomic investigation of the rabbit vitreous has identified a set of proteins that help further our understanding of the pathogenesis of rhegmatogenous retinal detachment and its complications.


docid: moattmgh

title: CD11c(hi) Dendritic Cells Regulate Ly-6C(hi) Monocyte Differentiation to Preserve Immune-privileged CNS in Lethal Neuroinflammation

text: Although the roles of dendritic cells (DCs) in adaptive defense have been defined well, the contribution of DCs to T cell-independent innate defense and subsequent neuroimmunopathology in immune-privileged CNS upon infection with neurotropic viruses has not been completely defined. Notably, DC roles in regulating innate CD11b(+)Ly-6C(hi) monocyte functions during neuroinflammation have not yet been addressed. Using selective ablation of CD11c(hi)PDCA-1(int/lo) DCs without alteration in CD11c(int)PDCA-1(hi) plasmacytoid DC number, we found that CD11c(hi) DCs are essential to control neuroinflammation caused by infection with neurotropic Japanese encephalitis virus, through early and increased infiltration of CD11b(+)Ly-6C(hi) monocytes and higher expression of CC chemokines. More interestingly, selective CD11c(hi) DC ablation provided altered differentiation and function of infiltrated CD11b(+)Ly-6C(hi) monocytes in the CNS through Flt3-L and GM-CSF, which was closely associated with severely enhanced neuroinflammation. Furthermore, CD11b(+)Ly-6C(hi) monocytes generated in CD11c(hi) DC-ablated environment had a deleterious rather than protective role during neuroinflammation, and were more quickly recruited into inflamed CNS, depending on CCR2, thereby exacerbating neuroinflammation via enhanced supply of virus from the periphery. Therefore, our data demonstrate that CD11c(hi) DCs provide a critical and unexpected role to preserve the immune-privileged CNS in lethal neuroinflammation via regulating the differentiation, function, and trafficking of CD11b(+)Ly-6C(hi) monocytes.


title: Loss of CARD9-mediated innate activation attenuates severe influenza pneumonia without compromising host viral immunity

text: Influenza virus (IFV) infection is a common cause of severe viral pneumonia associated with acute respiratory distress syndrome (ARDS), which is difficult to control with general immunosuppressive therapy including corticosteroids due to the unfavorable effect on viral replication. Studies have suggested that the excessive activation of the innate immunity by IFV is responsible for severe pathologies. In this study, we focused on CARD9, a signaling adaptor known to regulate innate immune activation through multiple innate sensor proteins, and investigated its role in anti-IFV defense and lung pathogenesis in a mouse model recapitulating severe influenza pneumonia with ARDS. We found that influenza pneumonia was dramatically attenuated in Card9-deficient mice, which showed improved mortality with reduced inflammatory cytokines and chemokines in the infected lungs. However, viral clearance, type-I interferon production, and the development of anti-viral B and T cell immunity were not compromised by CARD9 deficiency. Syk or CARD9-deficient DCs but not macrophages showed impaired cytokine but not type-I interferon production in response to IFV in vitro, indicating a possible role for the Syk-CARD9 pathway in DCs in excessive inflammation of IFV-infected lungs. Therefore, inhibition of this pathway is an ideal therapeutic target for severe influenza pneumonia without affecting viral clearance.


docid: xlrnhmon

title: Protection against Foot-and-Mouth Disease Virus in Guinea Pigs via Oral Administration of Recombinant Lactobacillus plantarum Expressing VP1

text: Mucosal vaccination is an effective strategy for generating antigen-specific immune responses against mucosal infections of foot-and-mouth disease virus (FMDV). In this study, Lactobacillus plantarum strains NC8 and WCFS1 were used as oral delivery vehicles containing a pSIP411-VP1 recombinant plasmid to initiate mucosal and systemic immune responses in guinea pigs. Guinea pigs were orally vaccinated (three doses) with NC8-pSIP411, NC8-pSIP411-VP1, WCFS1-pSIP411, WCFS1-pSIP411-VP1 or milk. Animals immunized with NC8-pSIP411-VP1 and WCFS1-pSIP411-VP1 developed high levels of antigen-specific serum IgG, IgA, IgM, mucosal secretory IgA (sIgA) and neutralizing antibodies, and revealed stronger cell-mediated immune responses and enhanced protection against FMDV challenge compared with control groups. The recombinant pSIP411-VP1 effectively improved immunoprotection against FMDV in guinea pigs.


title: Infectious Progeny of 2009 A (H1N1) Influenza Virus Replicated in and Released from Human Neutrophils

text: Various reports have indicated that a number of viruses could infect neutrophils, but the multiplication of viruses in neutrophils was abortive. Based on our previous finding that avian influenza viral RNA and proteins were present in the nucleus of infected human neutrophils in vivo, we investigated the possibility of 2009 A (H1N1) influenza viral synthesis in infected neutrophils and possible release of infectious progeny from host cells. In this study we found that human neutrophils in vitro without detectable level of sialic acid expression could be infected by this virus strain. We also show that the infected neutrophils can not only synthesize 2009 A (H1N1) viral mRNA and proteins, but also produce infectious progeny. These findings suggest that infectious progeny of 2009 A (H1N1) influenza virus could be replicated in and released from human neutrophils with possible clinical implications.


title: Salmonella enterica Serovars Enteritidis Infection Alters the Indigenous Microbiota Diversity in Young Layer Chicks

text: Avian gastrointestinal (GI) tracts are highly populated with a diverse array of microorganisms that share a symbiotic relationship with their hosts and contribute to the overall health and disease state of the intestinal tract. The microbiome of the young chick is easily prone to alteration in its composition by both exogenous and endogenous factors, especially during the early posthatch period. The genetic background of the host and exposure to pathogens can impact the diversity of the microbial profile that consequently contributes to the disease progression in the host. The objective of this study was to profile the composition and structure of the gut microbiota in young chickens from two genetically distinct highly inbred lines. Furthermore, the effect of the Salmonella Enteritidis infection on altering the composition makeup of the chicken microbiome was evaluated through the 16S rRNA gene sequencing analysis. One-day-old layer chicks were challenged with S. Enteritidis and the host cecal microbiota profile as well as the degree of susceptibility to Salmonella infection was examined at 2 and 7 days post infection. Our result indicated that host genotype had a limited effect on resistance to S. Enteritidis infection. Alpha diversity, beta diversity, and overall microbiota composition were analyzed for four factors: host genotype, age, treatment, and postinfection time points. S. Enteritidis infection in young chicks was found to significantly reduce the overall diversity of the microbiota population with expansion of Enterobacteriaceae family. These changes indicated that Salmonella colonization in the GI tract of the chickens has a direct effect on altering the natural development of the GI microbiota. The impact of S. Enteritidis infection on microbial communities was also more substantial in the late stage of infection. Significant inverse correlation between Enterobacteriaceae and Lachnospiraceae family in both non-infected and infected groups, suggested possible antagonistic interaction between members of these two taxa, which could potentially influences the overall microbial population in the gut. Our results also revealed that genetic difference between two lines had minimal effect on the establishment of microbiota population. Overall, this study provided preliminary insights into the contributing role of S. Enteritidis in influencing the overall makeup of chicken’s gut microbiota.


title: Porcine Epidemic Diarrhea Virus among Farmed Pigs, Ukraine

text: An outbreak of porcine epidemic diarrhea occurred in the summer of 2014 in Ukraine, severely affecting piglets <10 days of age; the mortality rate approached 100%. Full genome sequencing showed the virus to be closely related to strains reported from North America, showing a sequence identity of up to 99.8%.


title: No Evidence of Gouléako and Herbert Virus Infections in Pigs, Côte d’Ivoire and Ghana

text: A recent report suggested that 2 novel bunyaviruses discovered in insects in Côte d’Ivoire caused lethal disease in swine in South Korea. We conducted cell culture studies and tested serum from pigs exposed to mosquitoes in Côte d’Ivoire and Ghana and found no evidence for infection in pigs.


title: The baseline characteristics and interim analyses of the high-risk sentinel cohort of the Vietnam Initiative on Zoonotic InfectiONS (VIZIONS)

text: The Vietnam Initiative for Zoonotic Infections (VIZIONS) includes community-based ‘high-risk sentinel cohort’ (HRSC) studies investigating individuals at risk of zoonotic infection due to occupational or residential exposure to animals. A total of 852 HRSC members were recruited between March 2013 and August 2014 from three provinces (Ha Noi, Dak Lak, and Dong Thap). The most numerous group (72.8%) corresponded to individuals living on farms, followed by slaughterers (16.3%) and animal health workers (8.5%). Nasal/pharyngeal and rectal swabs were collected from HRSC members at recruitment and after notifying illness. Exposure to exotic animals (including wild pigs, porcupine, monkey, civet, bamboo rat and bat) was highest for the Dak Lak cohort (53.7%), followed by Ha Noi (13.7%) and Dong Thap (4.0%). A total of 26.8% of individuals reported consumption of raw blood over the previous year; 33.6% slaughterers reported no use of protective equipment at work. Over 686 person-years of observation, 213 episodes of suspect infectious disease were notified, equivalent of 0.35 reports per person-year. Responsive samples were collected from animals in the farm cohort. There was noticeable time and space clustering of disease episodes suggesting that the VIZIONS set up is also suitable for the formal epidemiological investigation of disease outbreaks.


title: Identification of climate factors related to human infection with avian influenza A H7N9 and H5N1 viruses in China

text: Human influenza infections display a strongly seasonal pattern. However, whether H7N9 and H5N1 infections correlate with climate factors has not been examined. Here, we analyzed 350 cases of H7N9 infection and 47 cases of H5N1 infection. The spatial characteristics of these cases revealed that H5N1 infections mainly occurred in the South, Middle, and Northwest of China, while the occurrence of H7N9 was concentrated in coastal areas of East and South of China. Aside from spatial-temporal characteristics, the most adaptive meteorological conditions for the occurrence of human infections by these two viral subtypes were different. We found that H7N9 infections correlate with climate factors, especially temperature (TEM) and relative humidity (RHU), while H5N1 infections correlate with TEM and atmospheric pressure (PRS). Hence, we propose a risky window (TEM 4–14 °C and RHU 65–95%) for H7N9 infection and (TEM 2–22 °C and PRS 980-1025 kPa) for H5N1 infection. Our results represent the first step in determining the effects of climate factors on two different virus infections in China and provide warning guidelines for the future when provinces fall into the risky windows. These findings revealed integrated predictive meteorological factors rooted in statistic data that enable the establishment of preventive actions and precautionary measures against future outbreaks.


docid: skambgbi

title: Quantity and Size Distribution of Cough-Generated Aerosol Particles Produced by Influenza Patients During and After Illness

text: The question of whether influenza is transmitted to a significant degree by aerosols remains controversial, in part, because little is known about the quantity and size of potentially infectious airborne particles produced by people with influenza. In this study, the size and amount of aerosol particles produced by nine subjects during coughing were measured while they had influenza and after they had recovered, using a laser aerosol particle spectrometer with a size range of 0.35 to 10 μm. Individuals with influenza produce a significantly greater volume of aerosol when ill compared with afterward (p = 0.0143). When the patients had influenza, their average cough aerosol volume was 38.3 picoliters (pL) of particles per cough (SD 43.7); after patients recovered, the average volume was 26.4 pL per cough (SD 45.6). The number of particles produced per cough was also higher when subjects had influenza (average 75,400 particles/cough, SD 97,300) compared with afterward (average 52,200, SD 98,600), although the difference did not reach statistical significance (p = 0.1042). The average number of particles expelled per cough varied widely from patient to patient, ranging from 900 to 302,200 particles/cough while subjects had influenza and 1100 to 308,600 particles/cough after recovery. When the subjects had influenza, an average of 63% of each subject's cough aerosol particle volume in the detection range was in the respirable size fraction (SD 22%), indicating that these particles could reach the alveolar region of the lungs if inhaled by another person. This enhancement in aerosol generation during illness may play an important role in influenza transmission and suggests that a better understanding of this phenomenon is needed to predict the production and dissemination of influenza-laden aerosols by people infected with this virus. [Supplementary materials are available for this article. Go to the publisher's online edition of Journal of Occupational and Environmental Hygiene for the following free supplemental resources: a PDF file of demographic information, influenza test results, and volume and peak flow rate during each cough and a PDF file containing number and size of aerosol particles produced.]


title: Gene silencing of TACE enhances plaque stability and improves vascular remodeling in a rabbit model of atherosclerosis

text: We aimed to test the hypothesis that gene silencing of tumor necrosis factor alpha converting enzyme (TACE) may attenuate lesion inflammation and positive vascular remodeling and enhance plaque stability in a rabbit model of atherosclerosis. Lentivirus-mediated TACE shRNA was injected into the abdominal aortic plaques of rabbits which effectively down-regulated TACE expression and activities from week 8 to week 16. TACE gene silencing reduced remodeling index and plaque burden, and diminished the content of macrophages and lipids while increased that of smooth muscle cells and collagen in the aortic plaques. In addition, TACE gene silencing attenuated the local expression of P65, iNOS, ICAM-1, VEGF and Flt-1 and activities of MMP9 and MMP2 while increased the local expression of TGF-β1 together with reduced number of neovessels in the aorta. TACE shRNA treatment resulted in down-regulated expression of TACE in macrophages and blunted ERK-P38 phosphorylation and tube formation of co-cultured mouse vascular smooth muscle cells or human umbilical vein endothelial cells. In conclusion, gene silencing of TACE enhanced plaque stability and improved vascular positive remodeling. The mechanisms may involve attenuated local inflammation, neovascularization and MMP activation, as well as enhanced collagen production probably via down-regulated ERK-NF-κB and up-regulated TGF-β1 signaling pathways.


title: Identification of potential drug targets by subtractive genome analysis of Escherichia coli O157:H7: an in silico approach

text: Bacterial enteric infections resulting in diarrhea, dysentery, or enteric fever constitute a huge public health problem, with more than a billion episodes of disease annually in developing and developed countries. In this study, the deadly agent of hemorrhagic diarrhea and hemolytic uremic syndrome, Escherichia coli O157:H7 was investigated with extensive computational approaches aimed at identifying novel and broad-spectrum antibiotic targets. A systematic in silico workflow consisting of comparative genomics, metabolic pathways analysis, and additional drug prioritizing parameters was used to identify novel drug targets that were essential for the pathogen’s survival but absent in its human host. Comparative genomic analysis of Kyoto Encyclopedia of Genes and Genomes annotated metabolic pathways identified 350 putative target proteins in E. coli O157:H7 which showed no similarity to human proteins. Further bio-informatic approaches including prediction of subcellular localization, calculation of molecular weight, and web-based investigation of 3D structural characteristics greatly aided in filtering the potential drug targets from 350 to 120. Ultimately, 44 non-homologous essential proteins of E. coli O157:H7 were prioritized and proved to have the eligibility to become novel broad-spectrum antibiotic targets and DNA polymerase III alpha (dnaE) was the top-ranked among these targets. Moreover, druggability of each of the identified drug targets was evaluated by the DrugBank database. In addition, 3D structure of the dnaE was modeled and explored further for in silico docking with ligands having potential druggability. Finally, we confirmed that the compounds N-coeleneterazine and N-(1,4-dihydro-5H-tetrazol-5-ylidene)-9-oxo-9H-xanthene-2-sulfon-amide were the most suitable ligands of dnaE and hence proposed as the potential inhibitors of this target protein. The results of this study could facilitate the discovery and release of new and effective drugs against E. coli O157:H7 and other deadly human bacterial pathogens.


title: Survival in Idiopathic pulmonary fibrosis acute exacerbations: the non-steroid approach

text: BACKGROUND: Idiopathic pulmonary fibrosis acute exacerbation (IPF-AE) constitutes IPF’s most devastating event, representing the unexpected superimposition of diffuse alveolar damage of unknown etiology. Guidelines recommend high-dose steroids treatment despite unproven benefit. We hypothesized that previous immunosuppression and the administration of high-dose steroids adversely affect IPF-AE outcome. METHODS: We studied all consecutive patients hospitalized in our department for IPF deterioration from 2007 to June 2013. Our protocol consisted of immediate cessation of immunosuppression (if any), best supportive care, broad-spectrum antimicrobials and thorough evaluation to detect reversible causes of deterioration. Patients were followed-up for survival; post-discharge none received immunosuppression. RESULTS: Twenty-four out of 85 admissions (28 %) fulfilled IPF-AE criteria. IPF-AE were analyzed both as unique events and as unique patients. As unique events 50 % survived; 3 out of 12 (25 %) in the group previously treated with immunosuppression whereas nine out of 12 (75 %) in the group not receiving immunosuppression (p = 0.041). As unique patients 35.3 % survived; 3 out of 6 (50 %) in the never treated group whereas three out of 11 (27.3 %) in the group receiving immunosuppression (p = 0.685). The history of immunosuppression significantly and adversely influenced survival (p = 0.035). Survival was greater in the never treated group compared to the immunosuppressed patients (p = 0.022). Post-discharge, our IPF-AE survivors had an 83 % 1-year survival. CONCLUSIONS: By applying the above mentioned protocol half of our patients survived. The history of immunosuppression before IPF-AE adversely influences survival. Avoiding steroids in IPF patients may favor the natural history of the disease even at the moment of its most devastating event.


docid: qutbqcwb

title: CVTree3 Web Server for Whole-genome-based and Alignment-free Prokaryotic Phylogeny and Taxonomy

text: A faithful phylogeny and an objective taxonomy for prokaryotes should agree with each other and ultimately follow the genome data. With the number of sequenced genomes reaching tens of thousands, both tree inference and detailed comparison with taxonomy are great challenges. We now provide one solution in the latest Release 3.0 of the alignment-free and whole-genome-based web server CVTree3. The server resides in a cluster of 64 cores and is equipped with an interactive, collapsible, and expandable tree display. It is capable of comparing the tree branching order with prokaryotic classification at all taxonomic ranks from domains down to species and strains. CVTree3 allows for inquiry by taxon names and trial on lineage modifications. In addition, it reports a summary of monophyletic and non-monophyletic taxa at all ranks as well as produces print-quality subtree figures. After giving an overview of retrospective verification of the CVTree approach, the power of the new server is described for the mega-classification of prokaryotes and determination of taxonomic placement of some newly-sequenced genomes. A few discrepancies between CVTree and 16S rRNA analyses are also summarized with regard to possible taxonomic revisions. CVTree3 is freely accessible to all users at http://tlife.fudan.edu.cn/cvtree3/ without login requirements.


title: TGF-βI Regulates Cell Migration through Pluripotent Transcription Factor OCT4 in Endometriosis

text: Transforming growth factor (TGF-β)/TGF-β receptor signal is known to promote cell migration. Up-regulation of TGF-β in serum/peritoneal fluid and increased levels of pluripotent transcription factor OCT4 in endometriotic tissues are frequently observed in patients with endometriosis. However, the mechanisms underlying how TGF-β/TGF-β receptor and OCT4 affect endometriotic cell migration still remain largely unknown. Therefore, endometriotic tissue with high cell migratory capacity were collected from patients with adenomyotic myometrium (n = 23) and chocolate cyst (n = 24); and endometrial tissue with low cell migratory capacity in normal endometrium or hyperplastic endometrium (n = 8) were collected as the controls. We found the mRNA levels of TGF-β receptor I (TGF-β RI) and OCT4 were significantly higher in the high-migratory ectopic endometriotic tissues than those of the low-migratory normal or hyperplastic endometrium. Positive correlations between TGF-β RI and OCT4, and either TGF-β RI or OCT4 with migration-related genes (SNAIL, SLUG and TWIST) regarding the mRNA levels were observed in human endometriotic tissues. TGF-βI dose-dependently increased the gene and protein levels of OCT4, SNAIL and N-Cadherin (N-CAD) and silencing of endogenous OCT4 significantly suppressed the TGF-βI-induced expressions of N-CAD and SNAIL in primary human endometriotic stromal cells and human endometrial carcinoma cell lines RL95-2 and HEC1A. Furthermore, TGF-βI significantly increased the migration ability of endometriotic cells and silencing of OCT4 dramatically suppressed the TGF-βI-induced cell migration activity evidenced by wound-closure assay, transwell assay, and confocal image of F-actin cellular distribution. In conclusion, the present findings demonstrate that the niche TGF-β plays a critical role in initiating expressions of pluripotent transcription factor OCT4 which may contribute to the ectopic endometrial growth by stimulating endometrial cell migration. These findings would be useful for developing therapeutic strategies targeting TGF-β-OCT4 signaling to prevent endometriosis in the future.


title: Complete Genome Sequence of the Porcine Epidemic Diarrhea Virus Variant CH/HNYF/2014

text: Sow’s milk is a potential route for the vertical transmission of porcine epidemic diarrhea virus (PEDV) from sow to suckling piglet. We report here the complete genome sequence of PEDV strain CH/HNYF/2014, which was isolated from milk samples. This information provides further understanding of the transmission mechanisms and genetic diversity of PEDV.


title: Expression of arginase I and inducible nitric oxide synthase in the peripheral blood and lymph nodes of HIV-positive patients

text: Arginase I (Arg I) and inducible nitric oxide synthase (iNOS) are important in regulating immune functions through their metabolites. Previous studies have revealed that the expression of Arg I is increased and the expression of iNOS is reduced in the serum and peripheral blood mononuclear cells of human immunodeficiency virus (HIV)-infected patients. As one of the most important immune organs and HIV replication sites, whether similar changes are present in the lymph nodes following HIV infection remains to be elucidated. To investigate this, the present study collected lymph node and blood specimens from 52 HIV-infected patients to measure the expression levels of Arg I and iNOS by immunohistochemistry and fluoresence-based flow cytometry. Compared with control subjects without HIV infection, the patients with HIV had significantly higher expression levels of Arg I in the lymph nodes and higher frequencies of Arg I(+) CD4(+) T cells and CD8(+) T cells in the blood and lymph nodes, and these results were contrary the those of iNOS in the corresponding compartments. The expression levels of Arg I in the lymph nodes and blood were negatively associated with peripheral CD4(+) T cell count and positively associated with viral load. However, the expression levels of iNOS in the lymph nodes and blood were positively associated with peripheral CD4(+) T cell count and negatively associated with viral load. These results showed that alterations in the expression levels of Arg I and iNOS in the peripheral T cells and peripheral nodes of HIV infected patients are associated with disease progression in these patients. These results indicate a potential to therapeutic strategy for delaying disease progression through regulating and manipulating the expression levels of Arg I and iNOS in patients infected with HIV.


title: Strategies for Pharmacological Organoprotection during Extracorporeal Circulation Targeting Ischemia-Reperfusion Injury

text: Surgical correction of congenital cardiac malformations or aortocoronary bypass surgery in many cases implies the use of cardiopulmonary-bypass (CPB). However, a possible negative impact of CPB on internal organs such as brain, kidney, lung and liver cannot be neglected. In general, CPB initiates a systemic inflammatory response (SIRS) which is presumably caused by contact of blood components with the surface of CPB tubing. Moreover, during CPB the heart typically undergoes a period of cold ischemia, and the other peripheral organs a global low flow hypoperfusion. As a result, a plethora of pro-inflammatory mediators and cytokines is released activating different biochemical pathways, which finally may result in the occurrence of microthrombosis, microemboli, in depletion of coagulation factors and haemorrhagic diathesis besides typical ischemia-reperfusion injuries. In our review we will focus on possible pharmacological interventions in patients to decrease negative effects of CPB and to improve post-operative outcome with regard to heart and other organs like brain, kidney, or lung.


title: Gliopathy of Demyelinating and Non-Demyelinating Strains of Mouse Hepatitis Virus

text: Demyelination in the central nervous system induced by neurovirulent strains of Mouse Hepatitis Virus (MHV) is mediated by the viral spike glycoprotein, but it is not clear whether the mechanism of this disease pathology involves direct viral infection of oligodendrocytes. Detailed studies of glial cell tropism of MHV are presented, demonstrating that direct MHV infection of oligodendrocytes differs between demyelinating (RSA59) and non-demyelinating (RSMHV2) viral strains both in vitro and in vivo. Our results indicate that direct injury of mature oligodendrocytes is an important mechanism of virus-induced demyelination. In vivo, RSA59 infection was identified in spinal cord gray and white matter, but infected oligodendrocytes were restricted to white matter. In contrast, RSMHV2 infection was restricted to gray matter neurons and was not localized to oligodendrocytes. In vitro, RSA59 can infect both oligodendrocyte precursors and differentiated oligodendrocytes, whereas RSMHV2 can infect oligodendrocyte precursors but not differentiated oligodendrocytes. Viral spreading through axonal means to white matter and release of the demyelinating strain MHV at the nerve end is critical for oligodendrocytes infection and subsequent demyelination. Understanding the mechanisms by which known viruses effect demyelination in this animal model has important therapeutic implications in the treatment of human demyelinating disease.


title: Risk Distribution of Human Infections with Avian Influenza H7N9 and H5N1 virus in China

text: It has been documented that the epidemiological characteristics of human infections with H7N9 differ significantly between H5N1. However, potential factors that may explain the different spatial distributions remain unexplored. We use boosted regression tree (BRT) models to explore the association of agro-ecological, environmental and meteorological variables with the occurrence of human cases of H7N9 and H5N1, and map the probabilities of occurrence of human cases. Live poultry markets, density of human, coverage of built-up land, relative humidity and precipitation were significant predictors for both. In addition, density of poultry, coverage of shrub and temperature played important roles for human H7N9 infection, whereas human H5N1 infection was associated with coverage of forest and water body. Based on the risks and distribution of ecological characteristics which may facilitate the circulation of the two viruses, we found Yangtze River Delta and Pearl River Delta, along with a few spots on the southeast coastline, to be the high risk areas for H7N9 and H5N1. Additional, H5N1 risk spots were identified in eastern Sichuan and southern Yunnan Provinces. Surveillance of the two viruses needs to be enhanced in these high risk areas to reduce the risk of future epidemics of avian influenza in China.


title: Flunarizine Prevents Hepatitis C Virus Membrane Fusion in a Genotype-dependent Manner by Targeting the Potential Fusion Peptide within E1

text: To explore mechanisms of hepatitis C virus (HCV) replication we screened a compound library including licensed drugs. Flunarizine, a diphenylmethylpiperazine used to treat migraine, inhibited HCV cell entry in vitro and in vivo in a genotype-dependent fashion. Analysis of mosaic viruses between susceptible and resistant strains revealed that E1 and E2 glycoproteins confer susceptibility to flunarizine. Time of addition experiments and single particle tracking of HCV demonstrated that flunarizine specifically prevents membrane fusion. Related phenothiazines and pimozide also inhibited HCV infection and preferentially targeted HCV genotype 2 viruses. However, phenothiazines and pimozide exhibited improved genotype coverage including the difficult to treat genotype 3. Flunarizine-resistant HCV carried mutations within the alleged fusion peptide and displayed cross-resistance to these compounds, indicating that these drugs have a common mode of action. Conclusion: These observations reveal novel details about HCV membrane fusion. Moreover, flunarizine and related compounds represent first-in-class HCV fusion inhibitors that merit consideration for repurposing as cost-effective component of HCV combination therapies.


title: Viral Infection at High Magnification: 3D Electron Microscopy Methods to Analyze the Architecture of Infected Cells

text: As obligate intracellular parasites, viruses need to hijack their cellular hosts and reprogram their machineries in order to replicate their genomes and produce new virions. For the direct visualization of the different steps of a viral life cycle (attachment, entry, replication, assembly and egress) electron microscopy (EM) methods are extremely helpful. While conventional EM has given important information about virus-host cell interactions, the development of three-dimensional EM (3D-EM) approaches provides unprecedented insights into how viruses remodel the intracellular architecture of the host cell. During the last years several 3D-EM methods have been developed. Here we will provide a description of the main approaches and examples of innovative applications.


title: Molecular Epidemiology of Human Rhinoviruses and Enteroviruses Highlights Their Diversity in Sub-Saharan Africa

text: Human rhinoviruses (HRVs) and enteroviruses (HEVs) belong to the Enterovirus genus and are the most frequent cause of infection worldwide, but data on their molecular epidemiology in Africa are scarce. To understand HRV and HEV molecular epidemiology in this setting, we enrolled febrile pediatric patients participating in a large prospective cohort assessing the causes of fever in Tanzanian children. Naso/oropharyngeal swabs were systematically collected and tested by real-time RT-PCR for HRV and HEV. Viruses from positive samples were sequenced and phylogenetic analyses were then applied to highlight the HRV and HEV types as well as recombinant or divergent strains. Thirty-eight percent (378/1005) of the enrolled children harboured an HRV or HEV infection. Although some types were predominant, many distinct types were co-circulating, including a vaccinal poliovirus, HEV-A71 and HEV-D68. Three HRV-A recombinants were identified: HRV-A36/HRV-A67, HRV-A12/HRV-A67 and HRV-A96/HRV-A61. Four divergent HRV strains were also identified: one HRV-B strain and three HRV-C strains. This is the first prospective study focused on HRV and HEV molecular epidemiology in sub-Saharan Africa. This systematic and thorough large screening with careful clinical data management confirms the wide genomic diversity of these viruses, brings new insights about their evolution and provides data about associated symptoms.


title: Recent Progress towards Novel EV71 Anti-Therapeutics and Vaccines

text: Enterovirus 71 (EV71) is a group of viruses that belongs to the Picornaviridae family, which also includes viruses such as polioviruses. EV71, together with coxsackieviruses, is widely known for its association with Hand Foot Mouth Disease (HFMD), which generally affects children age five and below. Besides HFMD, EV71 can also trigger more severe and life-threatening neurological conditions such as encephalitis. Considering the lack of a vaccine and antiviral drug against EV71, together with the increasing spread of these viruses, the development of such drugs and vaccines becomes the top priority in protecting our younger generations. This article, hence, reviews some of the recent progress in the formulations of anti-therapeutics and vaccine generation for EV71, covering (i) inactivated vaccines; (ii) baculovirus-expressed vaccines against EV71; (iii) human intravenous immunoglobulin (IVIg) treatment; and (iv) the use of monoclonal antibody therapy as a prevention and treatment for EV71 infections.


title: Two novel regulators of N‐acetyl‐galactosamine utilization pathway and distinct roles in bacterial infections

text: Bacterial pathogens can exploit metabolic pathways to facilitate their successful infection cycles, but little is known about roles of d‐galactosamine (GalN)/N‐acetyl‐d‐galactosamine (GalNAc) catabolism pathway in bacterial pathogenesis. Here, we report the genomic reconstruction of GalN/GalNAc utilization pathway in Streptococci and the diversified aga regulons. We delineated two new paralogous AgaR regulators for the GalN/GalNAc catabolism pathway. The electrophoretic mobility shift assays experiment demonstrated that AgaR2 (AgaR1) binds the predicted palindromes, and the combined in vivo data from reverse transcription quantitative polymerase chain reaction and RNA‐seq suggested that AgaR2 (not AgaR1) can effectively repress the transcription of the target genes. Removal of agaR2 (not agaR1) from Streptococcus suis 05ZYH33 augments significantly the abilities of both adherence to Hep‐2 cells and anti‐phagocytosis against RAW264.7 macrophage. As anticipated, the dysfunction in AgaR2‐mediated regulation of S. suis impairs its pathogenicity in experimental models of both mice and piglets. Our finding discovered two novel regulators specific for GalN/GalNAc catabolism and assigned them distinct roles into bacterial infections. To the best of our knowledge, it might represent a first paradigm that links the GalN/GalNAc catabolism pathway to bacterial pathogenesis.


title: A simple novel device for air sampling by electrokinetic capture

text: BACKGROUND: A variety of different sampling devices are currently available to acquire air samples for the study of the microbiome of the air. All have a degree of technical complexity that limits deployment. Here, we evaluate the use of a novel device, which has no technical complexity and is easily deployable. RESULTS: An air-cleaning device powered by electrokinetic propulsion has been adapted to provide a universal method for collecting samples of the aerobiome. Plasma-induced charge in aerosol particles causes propulsion to and capture on a counter-electrode. The flow of ions creates net bulk airflow, with no moving parts. A device and electrode assembly have been re-designed from air-cleaning technology to provide an average air flow of 120 lpm. This compares favorably with current air sampling devices based on physical air pumping. Capture efficiency was determined by comparison with a 0.4 μm polycarbonate reference filter, using fluorescent latex particles in a controlled environment chamber. Performance was compared with the same reference filter method in field studies in three different environments. For 23 common fungal species by quantitative polymerase chain reaction (qPCR), there was 100 % sensitivity and apparent specificity of 87 %, with the reference filter taken as “gold standard.” Further, bacterial analysis of 16S RNA by amplicon sequencing showed equivalent community structure captured by the electrokinetic device and the reference filter. Unlike other current air sampling methods, capture of particles is determined by charge and so is not controlled by particle mass. We analyzed particle sizes captured from air, without regard to specific analyte by atomic force microscopy: particles at least as low as 100 nM could be captured from ambient air. CONCLUSIONS: This work introduces a very simple plug-and-play device that can sample air at a high-volume flow rate with no moving parts and collect particles down to the sub-micron range. The performance of the device is substantially equivalent to capture by pumping through a filter for microbiome analysis by quantitative PCR and amplicon sequencing.


title: Live Cell Analysis and Mathematical Modeling Identify Determinants of Attenuation of Dengue Virus 2’-O-Methylation Mutant

text: Dengue virus (DENV) is the most common mosquito-transmitted virus infecting ~390 million people worldwide. In spite of this high medical relevance, neither a vaccine nor antiviral therapy is currently available. DENV elicits a strong interferon (IFN) response in infected cells, but at the same time actively counteracts IFN production and signaling. Although the kinetics of activation of this innate antiviral defense and the timing of viral counteraction critically determine the magnitude of infection and thus disease, quantitative and kinetic analyses are lacking and it remains poorly understood how DENV spreads in IFN-competent cell systems. To dissect the dynamics of replication versus antiviral defense at the single cell level, we generated a fully viable reporter DENV and host cells with authentic reporters for IFN-stimulated antiviral genes. We find that IFN controls DENV infection in a kinetically determined manner that at the single cell level is highly heterogeneous and stochastic. Even at high-dose, IFN does not fully protect all cells in the culture and, therefore, viral spread occurs even in the face of antiviral protection of naïve cells by IFN. By contrast, a vaccine candidate DENV mutant, which lacks 2’-O-methylation of viral RNA is profoundly attenuated in IFN-competent cells. Through mathematical modeling of time-resolved data and validation experiments we show that the primary determinant for attenuation is the accelerated kinetics of IFN production. This rapid induction triggered by mutant DENV precedes establishment of IFN-resistance in infected cells, thus causing a massive reduction of virus production rate. In contrast, accelerated protection of naïve cells by paracrine IFN action has negligible impact. In conclusion, these results show that attenuation of the 2’-O-methylation DENV mutant is primarily determined by kinetics of autocrine IFN action on infected cells.


title: Pre-Travel Medical Preparation of Business and Occupational Travelers: An Analysis of the Global TravEpiNet Consortium, 2009 to 2012

text: The aim of the study was to understand more about pre-travel preparations and itineraries of business and occupational travelers. METHODS: De-identified data from 18 Global TravEpiNet clinics from January 2009 to December 2012 were analyzed. RESULTS: Of 23,534 travelers, 61% were non-occupational and 39% occupational. Business travelers were more likely to be men, had short times to departure and shorter trip durations, and commonly refused influenza, meningococcal, and hepatitis B vaccines. Most business travelers indicated that employers suggested the pre-travel health consultation, whereas non-occupational travelers sought consultations because of travel health concerns. CONCLUSIONS: Sub-groups of occupational travelers have characteristic profiles, with business travelers being particularly distinct. Employers play a role in encouraging business travelers to seek pre-travel consultations. Such consultations, even if scheduled immediately before travel, can identify vaccination gaps and increase coverage.


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title: Polymicrobial intensive care unit-acquired pneumonia: prevalence, microbiology and outcome

text: BACKGROUND: Microbial aetiology of intensive care unit (ICU)-acquired pneumonia (ICUAP) determines antibiotic treatment and outcomes. The impact of polymicrobial ICUAP is not extensively known. We therefore investigated the characteristics and outcomes of polymicrobial aetiology of ICUAP. METHOD: Patients with ICUAP confirmed microbiologically were prospectively compared according to identification of 1 (monomicrobial) or more (polymicrobial) potentially-pathogenic microorganisms. Microbes usually considered as non-pathogenic were not considered for the etiologic diagnosis. We assessed clinical characteristics, microbiology, inflammatory biomarkers and outcome variables. RESULTS: Among 441 consecutive patients with ICUAP, 256 (58 %) had microbiologic confirmation, and 41 (16 %) of them polymicrobial pneumonia. Methicillin-sensitive Staphylococcus aureus, Haemophilus influenzae, and several Enterobacteriaceae were more frequent in polymicrobial pneumonia. Multi-drug and extensive-drug resistance was similarly frequent in both groups. Compared with monomicrobial, patients with polymicrobial pneumonia had less frequently chronic heart disease (6, 15 % vs. 71, 33 %, p = 0.019), and more frequently pleural effusion (18, 50 %, vs. 54, 25 %, p = 0.008), without any other significant difference. Appropriate empiric antimicrobial treatment was similarly frequent in the monomicrobial (185, 86 %) and the polymicrobial group (39, 95 %), as were the initial response to the empiric treatment, length of stay and mortality. Systemic inflammatory response was similar comparing monomicrobial with polymicrobial ICUAP. CONCLUSION: The aetiology of ICUAP confirmed microbiologically was polymicrobial in 16 % cases. Pleural effusion and absence of chronic heart disease are associated with polymicrobial pneumonia. When empiric treatment is frequently appropriate, polymicrobial aetiology does not influence the outcome of ICUAP.


title: Effects of Ultraviolet Germicidal Irradiation (UVGI) on N95 Respirator Filtration Performance and Structural Integrity

text: The ability to disinfect and reuse disposable N95 filtering facepiece respirators (FFRs) may be needed during a pandemic of an infectious respiratory disease such as influenza. Ultraviolet germicidal irradiation (UVGI) is one possible method for respirator disinfection. However, UV radiation degrades polymers, which presents the possibility that UVGI exposure could degrade the ability of a disposable respirator to protect the worker. To study this, we exposed both sides of material coupons and respirator straps from four models of N95 FFRs to UVGI doses from 120–950 J/cm(2). We then tested the particle penetration, flow resistance, and bursting strengths of the individual respirator coupon layers, and the breaking strength of the respirator straps. We found that UVGI exposure led to a small increase in particle penetration (up to 1.25%) and had little effect on the flow resistance. UVGI exposure had a more pronounced effect on the strengths of the respirator materials. At the higher UVGI doses, the strength of the layers of respirator material was substantially reduced (in some cases, by >90%). The changes in the strengths of the respirator materials varied considerably among the different models of respirators. UVGI had less of an effect on the respirator straps; a dose of 2360 J/cm(2) reduced the breaking strength of the straps by 20–51%. Our results suggest that UVGI could be used to effectively disinfect disposable respirators for reuse, but the maximum number of disinfection cycles will be limited by the respirator model and the UVGI dose required to inactivate the pathogen.


title: Ultrastructure and lipid composition of detergent-resistant membranes derived from mammalian sperm and two types of epithelial cells

text: Lipid rafts are micro-domains of ordered lipids (L(o) phase) in biological membranes. The L(o) phase of cellular membranes can be isolated from disordered lipids (L(d) phase) after treatment with 1 % Triton X-100 at 4 °C in which the L(o) phase forms the detergent-resistant membrane (DRM) fraction. The lipid composition of DRM derived from Madin-Darby canine kidney (MDCK) cells, McArdle cells and porcine sperm is compared with that of the whole cell. Remarkably, the unsaturation and chain length degree of aliphatic chains attached to phospholipids is virtually the same between DRM and whole cells. Cholesterol and sphingomyelin were enriched in DRMs but to a cell-specific molar ratio. Sulfatides (sphingolipids from MDCK cells) were enriched in the DRM while a seminolipid (an alkylacylglycerolipid from sperm) was depleted from the DRM. Treatment with <5 mM methyl-ß-cyclodextrin (MBCD) caused cholesterol removal from the DRM without affecting the composition and amount of the phospholipid while higher levels disrupted the DRM. The substantial amount of (poly)unsaturated phospholipids in DRMs as well as a low stoichiometric amount of cholesterol suggest that lipid rafts in biological membranes are more fluid and dynamic than previously anticipated. Using negative staining, ultrastructural features of DRM were monitored and in all three cell types the DRMs appeared as multi-lamellar vesicular structures with a similar morphology. The detergent resistance is a result of protein–cholesterol and sphingolipid interactions allowing a relatively passive attraction of phospholipids to maintain the L(o) phase. For this special issue, the relevance of our findings is discussed in a sperm physiological context.


title: Acute respiratory distress syndrome and acute myocarditis developed in a previously healthy adult with influenza B

text: BACKGROUND: Influenza B virus infection is generally considered to be mild and is rarely associated pulmonary cardiovascular involvement in adults. However fatal complications may occur. CASE PRESENTATION: A 43-year-old previously healthy Taiwanese male came to our emergency department due to high fever, chills, general malaise and myalgia for about 4 days. An influenza rapid test from a throat swab was negative. Chest radiography showed mild left lung infiltration and levofloxacin was prescribed. However, progressive shortness of breath and respiratory failure developed 48 h later after hospitalization. Emergent intubation was performed and he was transferred to the intensive care unit where oseltamivir (Tamiflu, Roche) 75 mg orally twice daily was given immediately. In the intensive care unit, cardiac catheterization revealed normal coronary arteries. However, a markedly elevated cardiac enzyme level (Troponin I level was up to 71.01 ng/ml), a positive cardiac magnetic resonance imaging findings and no coronary artery stenosis led to the diagnosis of acute myocarditis. Subsequent real-time polymerase chain reaction of endotracheal aspirates was positive for influenza B. His condition gradually improved and he was successfully weaned from the ventilator on day 22. He was discharged without prominent complications on day 35. CONCLUSION: Influenza B infection is not always a mild disease. Early detection, early administration of antiviral agents, appropriate antibiotics and best supportive care, is still the gold standard for patients such as the one reported.


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title: Lys-315 at the Interfaces of Diagonal Subunits of δ-Crystallin Plays a Critical Role in the Reversibility of Folding and Subunit Assembly

text: δ-Crystallin is the major structural protein in avian eye lenses and is homologous to the urea cycle enzyme argininosuccinate lyase. This protein is structurally assembled as double dimers. Lys-315 is the only residue which is arranged symmetrically at the diagonal subunit interfaces to interact with each other. This study found that wild-type protein had both dimers and monomers present in 2–4 M urea whilst only monomers of the K315A mutant were observed under the same conditions, as judged by sedimentation velocity analysis. The assembly of monomeric K315A mutant was reversible in contrast to wild-type protein. Molecular dynamics simulations showed that the dissociation of primary dimers is prior to the diagonal dimers in wild-type protein. These results suggest the critical role of Lys-315 in stabilization of the diagonal dimer structure. Guanidinium hydrochloride (GdmCl) denatured wild-type or K315A mutant protein did not fold into functional protein. However, the urea dissociated monomers of K315A mutant protein in GdmCl were reversible folding through a multiple steps mechanism as measured by tryptophan and ANS fluorescence. Two partly unfolded intermediates were detected in the pathway. Refolding of the intermediates resulted in a conformation with greater amounts of hydrophobic regions exposed which was prone to the formation of protein aggregates. The formation of aggregates was not prevented by the addition of α-crystallin. These results highlight that the conformational status of the monomers is critical for determining whether reversible oligomerization or aggregate formation occurs.


title: Elevated level of renal xanthine oxidase mRNA transcription after nephropathogenic infectious bronchitis virus infection in growing layers

text: To assess relationships between xanthine oxidase (XOD) and nephropathogenic infectious bronchitis virus (NIBV) infection, 240 growing layers (35 days old) were randomly divided into two groups (infected and control) of 120 chickens each. Each chicken in the control and infected group was intranasally inoculated with 0.2 mL sterile physiological saline and virus, respectively, after which serum antioxidant parameters and renal XOD mRNA expression in growing layers were evaluated at 8, 15 and 22 days post-inoculation (dpi). The results showed that serum glutathione peroxidase and superoxide dismutase activities in the infected group were significantly lower than in the control group at 8 and 15 dpi (p < 0.01), while serum malondialdehyde concentrations were significantly higher (p < 0.01). The serum uric acid was significantly higher than that of the control group at 15 dpi (p < 0.01). In addition, the kidney mRNA transcript level and serum activity of XOD in the infected group was significantly higher than that of the control group at 8, 15 and 22 dpi (p < 0.05). The results indicated that NIBV infection could cause the increases of renal XOD gene transcription and serum XOD activity, leading to hyperuricemia and reduction of antioxidants in the body.


title: Magnetic nanoparticle based purification and enzyme-linked immunosorbent assay using monoclonal antibody against enrofloxacin

text: Monoclonal anti-enrofloxacin antibody was prepared for a direct competitive enzyme-linked immunosorbent assay (ELISA) and purification system using monoclonal antibody (mAb) coupled magnetic nanoparticles (MNPs). The IC(50) values of the developed mAb for enrofloxacin (ENR), ciprofloxacin, difloxacin, sarafloxacin, pefloxacin, and norfloxacin were 5.0, 8.3, 9.7, 21.7, 36.0, and 63.7 ng/mL, respectively. The lowest detectable level of ENR was 0.7 ng/mL in the prepared ELISA system. To validate the developed ELISA in the food matrix, known amounts of ENR were spiked in meat and egg samples at 10, 20 and 30 ng/mL. Recoveries for ENR ranged from 72.9 to 113.16% with a coefficient of variation (CV) of 2.42 to 10.11%. The applicability of the mAb-MNP system was verified by testing the recoveries for ENR residue in three different matrices. Recoveries for ENR ranged from 75.16 to 86.36%, while the CV ranged from 5.08 to 11.53%. Overall, ENR-specific monoclonal antibody was prepared and developed for use in competitive to ELISAs for the detection of ENR in animal meat samples. Furthermore, we suggest that a purification system for ENR using mAb-coupled MNPs could be useful for determination of ENR residue in food.


title: Anti-inflammatory effects of indirubin derivatives on influenza A virus-infected human pulmonary microvascular endothelial cells

text: Influenza A virus (IAV) poses global threats to human health. Acute respiratory distress syndrome and multi-organ dysfunction are major complications in patients with severe influenza infection. This may be explained by the recent studies which highlighted the role of the pulmonary endothelium as the center of innate immune cells recruitment and excessive pro-inflammatory cytokines production. In this report, we examined the potential immunomodulatory effects of two indirubin derivatives, indirubin-3′-(2,3-dihydroxypropyl)-oximether (E804) and indirubin-3′-oxime (E231), on IAV (H9N2) infected-human pulmonary microvascular endothelial cells (HPMECs). Infection of H9N2 on HPMECs induced a high level of chemokines and cytokines production including IP-10, RANTES, IL-6, IFN-β and IFN-γ1. Post-treatment of E804 or E231 could significantly suppress the production of these cytokines. H9N2 infection rapidly triggered the activation of innate immunity through phosphorylation of signaling molecules including mitogen-activated protein kinases (MAPKs) and signal transducer and activator of transcription (STAT) proteins. Using specific inhibitors or small-interfering RNA, we confirmed that indirubin derivatives can suppress H9N2-induced cytokines production through MAPKs and STAT3 signaling pathways. These results underscore the immunomodulatory effects of indirubin derivatives on pulmonary endothelium and its therapeutic potential on IAV-infection.


title: Genome-Wide Gene Expression Analysis Identifies the Proto-oncogene Tyrosine-Protein Kinase Src as a Crucial Virulence Determinant of Infectious Laryngotracheitis Virus in Chicken Cells

text: Given the side effects of vaccination against infectious laryngotracheitis (ILT), novel strategies for ILT control and therapy are urgently needed. The modulation of host-virus interactions is a promising strategy to combat the virus; however, the interactions between the host and avian ILT herpesvirus (ILTV) are unclear. Using genome-wide transcriptome studies in combination with a bioinformatic analysis, we identified proto-oncogene tyrosine-protein kinase Src (Src) to be an important modulator of ILTV infection. Src controls the virulence of ILTV and is phosphorylated upon ILTV infection. Functional studies revealed that Src prolongs the survival of host cells by increasing the threshold of virus-induced cell death. Therefore, Src is essential for viral replication in vitro and in ovo but is not required for ILTV-induced cell death. Furthermore, our results identify a positive-feedback loop between Src and the tyrosine kinase focal adhesion kinase (FAK), which is necessary for the phosphorylation of either Src or FAK and is required for Src to modulate ILTV infection. To the best of our knowledge, we are the first to identify a key host regulator controlling host-ILTV interactions. We believe that our findings have revealed a new potential therapeutic target for ILT control and therapy. IMPORTANCE Despite the extensive administration of live attenuated vaccines starting from the mid-20th century and the administration of recombinant vaccines in recent years, infectious laryngotracheitis (ILT) outbreaks due to avian ILT herpesvirus (ILTV) occur worldwide annually. Presently, there are no drugs or control strategies that effectively treat ILT. Targeting of host-virus interactions is considered to be a promising strategy for controlling ILTV infections. However, little is known about the mechanisms governing host-ILTV interactions. The results from our study advance our understanding of host-ILTV interactions on a molecular level and provide experimental evidence that it is possible to control ILT via the manipulation of host-virus interactions.


title: Database resources of the National Center for Biotechnology Information

text: The National Center for Biotechnology Information (NCBI) provides a large suite of online resources for biological information and data, including the GenBank(®) nucleic acid sequence database and the PubMed database of citations and abstracts for published life science journals. Additional NCBI resources focus on literature (PubMed Central (PMC), Bookshelf and PubReader), health (ClinVar, dbGaP, dbMHC, the Genetic Testing Registry, HIV-1/Human Protein Interaction Database and MedGen), genomes (BioProject, Assembly, Genome, BioSample, dbSNP, dbVar, Epigenomics, the Map Viewer, Nucleotide, Probe, RefSeq, Sequence Read Archive, the Taxonomy Browser and the Trace Archive), genes (Gene, Gene Expression Omnibus (GEO), HomoloGene, PopSet and UniGene), proteins (Protein, the Conserved Domain Database (CDD), COBALT, Conserved Domain Architecture Retrieval Tool (CDART), the Molecular Modeling Database (MMDB) and Protein Clusters) and chemicals (Biosystems and the PubChem suite of small molecule databases). The Entrez system provides search and retrieval operations for most of these databases. Augmenting many of the web applications are custom implementations of the BLAST program optimized to search specialized datasets. All of these resources can be accessed through the NCBI home page at www.ncbi.nlm.nih.gov.


title: Anti-foot-and-mouth disease virus effects of Chinese herbal kombucha in vivo

text: The foot and mouth disease virus (FMDV) is sensitive to acids and can be inactivated by exposure to low pH conditions. Spraying animals at risk of infection with suspensions of acid-forming microorganisms has been identified as a potential strategy for preventing FMD. Kombucha is one of the most strongly acid-forming symbiotic probiotics and could thus be an effective agent with which to implement this strategy. Moreover, certain Chinese herbal extracts are known to have broad-spectrum antiviral effects. Chinese herbal kombucha can be prepared by fermenting Chinese herbal extracts with a kombucha culture. Previous studies demonstrated that Chinese herbal kombucha prepared in this way efficiently inhibits FMDV replication in vitro. To assess the inhibitory effects of Chinese herbal kombucha against FMDV in vitro, swine challenged by intramuscular injection with 1000 SID(50) of swine FMDV serotype O strain O/China/99 after treatment with Chinese herbal kombucha were partially protected against infection, as demonstrated by a lack of clinical symptoms and qRT-PCR analysis. In a large scale field trial, spraying cattle in an FMD outbreak zone with kombucha protected against infection. Chinese herbal kombucha may be a useful probiotic agent for managing FMD outbreaks.


title: Deletion of Dystrophin In-Frame Exon 5 Leads to a Severe Phenotype: Guidance for Exon Skipping Strategies

text: Duchenne and Becker muscular dystrophy severity depends upon the nature and location of the DMD gene lesion and generally correlates with the dystrophin open reading frame. However, there are striking exceptions where an in-frame genomic deletion leads to severe pathology or protein-truncating mutations (nonsense or frame-shifting indels) manifest as mild disease. Exceptions to the dystrophin reading frame rule are usually resolved after molecular diagnosis on muscle RNA. We report a moderate/severe Becker muscular dystrophy patient with an in-frame genomic deletion of DMD exon 5. This mutation has been reported by others as resulting in Duchenne or Intermediate muscular dystrophy, and the loss of this in-frame exon in one patient led to multiple splicing events, including omission of exon 6, that disrupts the open reading frame and is consistent with a severe phenotype. The patient described has a deletion of dystrophin exon 5 that does not compromise recognition of exon 6, and although the deletion does not disrupt the reading frame, his clinical presentation is more severe than would be expected for classical Becker muscular dystrophy. We suggest that the dystrophin isoform lacking the actin-binding sequence encoded by exon 5 is compromised, reflected by the phenotype resulting from induction of this dystrophin isoform in mouse muscle in vivo. Hence, exon skipping to address DMD-causing mutations within DMD exon 5 may not yield an isoform that confers marked clinical benefit. Additional studies will be required to determine whether multi-exon skipping strategies could yield more functional dystrophin isoforms, since some BMD patients with larger in-frame deletions in this region have been reported with mild phenotypes.


title: Pilot study of participant-collected nasal swabs for acute respiratory infections in a low-income, urban population

text: OBJECTIVE: To assess the feasibility and validity of unsupervised participant-collected nasal swabs to detect respiratory pathogens in a low-income, urban minority population. METHODS: This project was conducted as part of an ongoing community-based surveillance study in New York City to identify viral etiologies of acute respiratory infection. In January 2014, following sample collection by trained research assistants, participants with acute respiratory infection from 30 households subsequently collected and returned a self-collected/parent-collected nasal swab via mail. Self/parental swabs corresponding with positive reverse transcription polymerase chain reaction primary research samples were analyzed. RESULTS: Nearly all (96.8%, n=30/31) households agreed to participate; 100% reported returning the sample and 29 were received (median time: 8 days). Most (18; 62.1%) of the primary research samples were positive. For eight influenza-positive research samples, seven (87.5%) self-swabs were also positive. For ten other respiratory pathogen-positive research samples, eight (80.0%) self-swabs were positive. Sensitivity of self-swabs for any respiratory pathogen was 83.3% and 87.5% for influenza, and specificity for both was 100%. There was no relationship between level of education and concordance of results between positive research samples and their matching participant swab. CONCLUSION: In this pilot study, self-swabbing was feasible and valid in a low-income, urban minority population.


docid: hrfbygub

title: Heterologous Immunity between Adenoviruses and Hepatitis C Virus: A New Paradigm in HCV Immunity and Vaccines

text: Adenoviruses (Ad) are commonly used as vectors for gene therapy and/or vaccine delivery. Recombinant Ad vectors are being tested as vaccines for many pathogens. We have made a surprising observation that peptides derived from various hepatitis C virus (HCV) antigens contain extensive regions of homology with multiple adenovirus proteins, and conclusively demonstrate that adenovirus vector can induce robust, heterologous cellular and humoral immune responses against multiple HCV antigens. Intriguingly, the induction of this cross-reactive immunity leads to significant reduction of viral loads in a recombinant vaccinia-HCV virus infected mouse model, supporting their role in antiviral immunity against HCV. Healthy human subjects with Ad-specific pre-existing immunity demonstrated cross-reactive cellular and humoral immune responses against multiple HCV antigens. These findings reveal the potential of a previously uncharacterized property of natural human adenovirus infection to dictate, modulate and/or alter the course of HCV infection upon exposure. This intrinsic property of adenovirus vectors to cross-prime HCV immunity can also be exploited to develop a prophylactic and/or therapeutic vaccine against HCV.


title: Real-Time Reverse Transcription PCR Assay for Detection of Senecavirus A in Swine Vesicular Diagnostic Specimens

text: Senecavirus A (SV-A), formerly, Seneca Valley virus (SVV), has been detected in swine with vesicular lesions and is thought to be associated with swine idiopathic vesicular disease (SIVD), a vesicular disease syndrome that lacks a defined causative agent. The clinical presentation of SIVD resembles that of other more contagious and economically devastating vesicular diseases, such as foot-and-mouth disease (FMD), swine vesicular disease (SVD), and vesicular stomatitis (VS), that typically require immediate rule out diagnostics to lift restrictions on animal quarantine, movement, and trade. This study presents the development of a sensitive, SYBR Green RT-qPCR assay suitable for detection of SV-A in diagnostic swine specimens. After testing 50 pigs with clinical signs consistent with vesicular disease, 44 (88%) were found to be positive for SV-A by RT-qPCR as compared to none from a negative cohort of 35 animals without vesicular disease, indicating that the assay is able to successfully detect the virus in an endemic population. SV-A RNA was also detectable at a low level in sera from a subset of pigs that presented with (18%) or without (6%) vesicular signs. In 2015, there has been an increase in the occurrence of SV-A in the US, and over 200 specimens submitted to our laboratory for vesicular investigation have tested positive for the virus using this method. SV-A RNA was detectable in all common types of vesicular specimens including swabs and tissue from hoof lesions, oral and snout epithelium, oral swabs, scabs, and internal organ tissues such as liver and lymph node. Genome sequencing analysis from recent virus isolates was performed to confirm target amplicon specificity and was aligned to previous isolates.


title: Statistical Evaluation of HTS Assays for Enzymatic Hydrolysis of β-Keto Esters

text: β-keto esters are used as precursors for the synthesis of β-amino acids, which are building blocks for some classes of pharmaceuticals. Here we describe the comparison of screening procedures for hydrolases to be used for the hydrolysis of β-keto esters, the first step in the preparation of β-amino acids. Two of the tested high throughput screening (HTS) assays depend on coupled enzymatic reactions which detect the alcohol released during ester hydrolysis by luminescence or absorption. The third assay detects the pH shift due to acid formation using an indicator dye. To choose the most efficient approach for screening, we assessed these assays with different statistical methods—namely, the classical Z’-factor, standardized mean difference (SSMD), the Kolmogorov-Smirnov-test, and t-statistics. This revealed that all three assays are suitable for HTS, the pH assay performing best. Based on our data we discuss the explanatory power of different statistical measures. Finally, we successfully employed the pH assay to identify a very fast hydrolase in an enzyme-substrate screening.


title: Abdominal Muscle Activity during Mechanical Ventilation Increases Lung Injury in Severe Acute Respiratory Distress Syndrome

text: OBJECTIVE: It has proved that muscle paralysis was more protective for injured lung in severe acute respiratory distress syndrome (ARDS), but the precise mechanism is not clear. The purpose of this study was to test the hypothesis that abdominal muscle activity during mechanically ventilation increases lung injury in severe ARDS. METHODS: Eighteen male Beagles were studied under mechanical ventilation with anesthesia. Severe ARDS was induced by repetitive oleic acid infusion. After lung injury, Beagles were randomly assigned into spontaneous breathing group (BIPAP(SB)) and abdominal muscle paralysis group (BIPAP(AP)). All groups were ventilated with BIPAP model for 8h, and the high pressure titrated to reached a tidal volume of 6ml/kg, the low pressure was set at 10 cmH(2)O, with I:E ratio 1:1, and respiratory rate adjusted to a PaCO(2) of 35–60 mmHg. Six Beagles without ventilator support comprised the control group. Respiratory variables, end-expiratory volume (EELV) and gas exchange were assessed during mechanical ventilation. The levels of Interleukin (IL)-6, IL-8 in lung tissue and plasma were measured by qRT-PCR and ELISA respectively. Lung injury scores were determined at end of the experiment. RESULTS: For the comparable ventilator setting, as compared with BIPAP(SB) group, the BIPAP(AP) group presented higher EELV (427±47 vs. 366±38 ml) and oxygenation index (293±36 vs. 226±31 mmHg), lower levels of IL-6(216.6±48.0 vs. 297.5±71.2 pg/ml) and IL-8(246.8±78.2 vs. 357.5±69.3 pg/ml) in plasma, and lower express levels of IL-6 mRNA (15.0±3.8 vs. 21.2±3.7) and IL-8 mRNA (18.9±6.8 vs. 29.5±7.9) in lung tissues. In addition, less lung histopathology injury were revealed in the BIPAP(AP) group (22.5±2.0 vs. 25.2±2.1). CONCLUSION: Abdominal muscle activity during mechanically ventilation is one of the injurious factors in severe ARDS, so abdominal muscle paralysis might be an effective strategy to minimize ventilator-induce lung injury.


title: Loss of cysteinyl-tRNA synthetase (CARS) induces the transsulfuration pathway and inhibits ferroptosis induced by cystine deprivation

text: Ferroptosis is a form of regulated non-apoptotic cell death that has been implicated in several disease contexts. A better understanding of the ferroptotic death mechanism could lead to the development of new therapeutics for degenerative diseases, and a better understanding of how to induce ferroptosis in specific tumor contexts. We performed an unbiased genome-wide siRNA screen to find genetic suppressors of ferroptosis. We determined that loss of CARS, the cysteinyl-tRNA synthetase, suppresses ferroptosis induced by erastin, which inhibits the cystine–glutamate antiporter known as system x(c)(−). Knockdown of CARS inhibited erastin-induced death by preventing the induction of lipid reactive oxygen species, without altering iron homeostasis. Knockdown of CARS led to the accumulation of cystathionine, a metabolite on the transsulfuration pathway, and upregulated genes associated with serine biosynthesis and transsulfuration. In addition, inhibition of the transsulfuration pathway resensitized cells to erastin, even after CARS knockdown. These studies demonstrate a new mechanism of resistance to ferroptosis and may lead to strategies for inducing and suppressing ferroptosis in diverse contexts.


title: Genome-wide Analysis of Epstein-Barr Virus (EBV) Integration and Strain in C666-1 and Raji Cells

text: EBV is a key risk factor for many malignancy diseases such as nasopharyngeal carcinoma (NPC) and Burkitt lymphoma (BL). EBV integration has been reported, but its scale and impact to cancer development is remains unclear. C666-1 (NPC cell line) and Raji (BL cell line) are commonly studied EBV-positive cancer cells. A rare few EBV integration sites in Raji were found in previous research by traditional methods. To deeply survey EBV integration, we sequenced C666-1 and Raji whole genomes by the next generation sequencing (NGS) technology and a total of 909 breakpoints were detected in the two cell lines. Moreover, we observed that the number of integration sites was positive correlated with the total amount of chromosome structural variations (SVs) and copy number structural variations (CNVs), and most breakpoints located inside or nearby genome structural variations regions. It suggested that host genome instability provided an opportunity for EBV integration on one hand and the integration aggravated host genome instability on the other hand. Then, we respectively assembled the C666-1 and Raji EBV strains which would be useful resources for EBV-relative studies. Thus, we report the most comprehensive characterization of EBV integration in NPC cell and BL cell, and EBV shows the wide range and random integration to increase the tumorigenesis. The NGS provides an incomparable level of resolution on EBV integration and a convenient approach to obtain viral strain compared to any research technology before.


title: Recent advances in biosynthesis of bioactive compounds in traditional Chinese medicinal plants

text: Plants synthesize and accumulate large amount of specialized (or secondary) metabolites also known as natural products, which provide a rich source for modern pharmacy. In China, plants have been used in traditional medicine for thousands of years. Recent development of molecular biology, genomics and functional genomics as well as high-throughput analytical chemical technologies has greatly promoted the research on medicinal plants. In this article, we review recent advances in the elucidation of biosynthesis of specialized metabolites in medicinal plants, including phenylpropanoids, terpenoids and alkaloids. These natural products may share a common upstream pathway to form a limited numbers of common precursors, but are characteristic in distinct modifications leading to highly variable structures. Although this review is focused on traditional Chinese medicine, other plants with a great medicinal interest or potential are also discussed. Understanding of their biosynthesis processes is critical for producing these highly value molecules at large scale and low cost in microbes and will benefit to not only human health but also plant resource conservation.


title: Endocytic function is critical for influenza A virus infection via DC-SIGN and L-SIGN

text: The ubiquitous presence of cell-surface sialic acid (SIA) has complicated efforts to identify specific transmembrane glycoproteins that function as bone fide entry receptors for influenza A virus (IAV) infection. The C-type lectin receptors (CLRs) DC-SIGN (CD209) and L-SIGN (CD209L) enhance IAV infection however it is not known if they act as attachment factors, passing virions to other unknown receptors for virus entry, or as authentic entry receptors for CLR-mediated virus uptake and infection. Sialic acid-deficient Lec2 Chinese Hamster Ovary (CHO) cell lines were resistant to IAV infection whereas expression of DC-SIGN/L-SIGN restored susceptibility of Lec2 cells to pH- and dynamin-dependent infection. Moreover, Lec2 cells expressing endocytosis-defective DC-SIGN/L-SIGN retained capacity to bind IAV but showed reduced susceptibility to infection. These studies confirm that DC-SIGN and L-SIGN are authentic endocytic receptors for IAV entry and infection.


title: A CRISPR/Cas9 and Cre/Lox system-based express vaccine development strategy against re-emerging Pseudorabies virus

text: Virus evolves rapidly to escape vaccine-induced immunity, posing a desperate demand for efficient vaccine development biotechnologies. Here we present an express vaccine development strategy based on CRISPR/Cas9 and Cre/Lox system against re-emerging Pseudorabies virus, which caused the recent devastating swine pseudorabies outbreak in China. By CRISPR/Cas9 system, the virulent genes of the newly isolated strain were simultaneously substituted by marker genes, which were subsequently excised using Cre/Lox system for vaccine safety concern. Notably, single cell FACS technology was applied to further promote virus purification efficiency. The combination of these state-of-art technologies greatly accelerated vaccine development. Finally, vaccination and challenge experiments proved this vaccine candidate’s protective efficacy in pigs and the promise to control current pseudorabies outbreak. This is, to our knowledge, the first successful vaccine development based on gene edit technologies, demonstrating these technologies leap from laboratory to industry. It may pave the way for future express antiviral vaccine development.


title: PB2-588 V promotes the mammalian adaptation of H10N8, H7N9 and H9N2 avian influenza viruses

text: Human infections with avian influenza H7N9 or H10N8 viruses have been reported in China, raising concerns that they might cause human epidemics and pandemics. However, how these viruses adapt to mammalian hosts is unclear. Here we show that besides the commonly recognized viral polymerase subunit PB2 residue 627 K, other residues including 87E, 292 V, 340 K, 588 V, 648 V, and 676 M in PB2 also play critical roles in mammalian adaptation of the H10N8 virus. The avian-origin H10N8, H7N9, and H9N2 viruses harboring PB2-588 V exhibited higher polymerase activity, more efficient replication in mammalian and avian cells, and higher virulence in mice when compared to viruses with PB2-588 A. Analyses of available PB2 sequences showed that the proportion of avian H9N2 or human H7N9 influenza isolates bearing PB2-588 V has increased significantly since 2013. Taken together, our results suggest that the substitution PB2-A588V may be a new strategy for an avian influenza virus to adapt mammalian hosts.


title: Rhinoviruses and Respiratory Enteroviruses: Not as Simple as ABC

text: Rhinoviruses (RVs) and respiratory enteroviruses (EVs) are leading causes of upper respiratory tract infections and among the most frequent infectious agents in humans worldwide. Both are classified in the Enterovirus genus within the Picornaviridae family and they have been assigned to seven distinct species, RV-A, B, C and EV-A, B, C, D. As viral infections of public health significance, they represent an important financial burden on health systems worldwide. However, the lack of efficient antiviral treatment or vaccines against these highly prevalent pathogens prevents an effective management of RV-related diseases. Current advances in molecular diagnostic techniques have revealed the presence of RV in the lower respiratory tract and its role in lower airway diseases is increasingly reported. In addition to an established etiological role in the common cold, these viruses demonstrate an unexpected capacity to spread to other body sites under certain conditions. Some of these viruses have received particular attention recently, such as EV-D68 that caused a large outbreak of respiratory illness in 2014, respiratory EVs from species C, or viruses within the newly-discovered RV-C species. This review provides an update of the latest findings on clinical and fundamental aspects of RV and respiratory EV, including a summary of basic knowledge of their biology.


title: Molecular Mechanisms of White Spot Syndrome Virus Infection and Perspectives on Treatments

text: Since its emergence in the 1990s, White Spot Disease (WSD) has had major economic and societal impact in the crustacean aquaculture sector. Over the years shrimp farming alone has experienced billion dollar losses through WSD. The disease is caused by the White Spot Syndrome Virus (WSSV), a large dsDNA virus and the only member of the Nimaviridae family. Susceptibility to WSSV in a wide range of crustacean hosts makes it a major risk factor in the translocation of live animals and in commodity products. Currently there are no effective treatments for this disease. Understanding the molecular basis of disease processes has contributed significantly to the treatment of many human and animal pathogens, and with a similar aim considerable efforts have been directed towards understanding host–pathogen molecular interactions for WSD. Work on the molecular mechanisms of pathogenesis in aquatic crustaceans has been restricted by a lack of sequenced and annotated genomes for host species. Nevertheless, some of the key host–pathogen interactions have been established: between viral envelope proteins and host cell receptors at initiation of infection, involvement of various immune system pathways in response to WSSV, and the roles of various host and virus miRNAs in mitigation or progression of disease. Despite these advances, many fundamental knowledge gaps remain; for example, the roles of the majority of WSSV proteins are still unknown. In this review we assess current knowledge of how WSSV infects and replicates in its host, and critique strategies for WSD treatment.


title: Sequence-based approach for rapid identification of cross-clade CD8+ T-cell vaccine candidates from all high-risk HPV strains

text: Human papilloma virus (HPV) is the primary etiological agent responsible for cervical cancer in women. Although in total 16 high-risk HPV strains have been identified so far. Currently available commercial vaccines are designed by targeting mainly HPV16 and HPV18 viral strains as these are the most common strains associated with cervical cancer. Because of the high level of antigenic specificity of HPV capsid antigens, the currently available vaccines are not suitable to provide cross-protection from all other high-risk HPV strains. Due to increasing reports of cervical cancer cases from other HPV high-risk strains other than HPV16 and 18, it is crucial to design vaccine that generate reasonable CD8+ T-cell responses for possibly all the high-risk strains. With this aim, we have developed a computational workflow to identify conserved cross-clade CD8+ T-cell HPV vaccine candidates by considering E1, E2, E6 and E7 proteins from all the high-risk HPV strains. We have identified a set of 14 immunogenic conserved peptide fragments that are supposed to provide protection against infection from any of the high-risk HPV strains across globe. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s13205-015-0352-z) contains supplementary material, which is available to authorized users.