Effective Elimination and Biodegradation of Polycyclic Aromatic Hydrocarbons from Seawater through the Formation of Magnetic Microfibres.

Supramolecular aggregates formed between polycyclic aromatic hydrocarbons and either naphthalene or perylene-derived diimides have been anchored in magnetite magnetic nanoparticles. The high affinity and stability of these aggregates allow them to capture and confine these extremely carcinogenic contaminants in a reduced space. In some cases, the high cohesion of these aggregates leads to the formation of magnetic microfibres of several microns in length, which can be isolated from the solution by the direct action of a magnet. Here we show a practical application of bioremediation aimed at the environmental decontamination of naphthalene, a very profuse contaminant, based on the uptake, sequestration, and acceleration of the biodegradation of the formed supramolecular aggregate, by the direct action of a bacterium of the lineage Roseobacter (biocompatible with nanostructured receptors and very widespread in marine environments) without providing more toxicity to the environment.

Genetic diversity of the 2013-14 human isolates of influenza H7N9 in China.

BACKGROUND: Influenza H7N9 has become an endemic pathogen in China where circulating virus is found extensively in wild birds and domestic poultry. Two epidemic waves of Human H7N9 infections have taken place in Eastern and South Central China during the years of 2013 and 2014. In this study, we report on the first four human cases of influenza H7N9 in Shantou, Guangdong province, which occurred during the second H7N9 wave, and the subsequent analysis of the viral isolates. METHODS: Viral genomes were subjected to multisegment amplification and sequenced in an Illumina MiSeq. Later, phylogenetic analyses of influenza H7N9 viruses were performed to establish the evolutionary context of the disease in humans. RESULTS: The sequences of the isolates from Shantou have closer evolutionary proximity to the predominant Eastern H7N9 cluster (similar to A/Shanghai/1/2013 (H7N9)) than to the Southern H7N9 cluster (similar to A/Guangdong/1/2013 (H7N9)). CONCLUSIONS: Two distinct phylogenetic groups of influenza H7N9 circulate currently in China and cause infections in humans as a consequence of cross-species spillover from the avian disease. The Eastern cluster, which includes the four isolates from Shantou, presents a wide geographic distribution and overlaps with the more restricted area of circulation of the Southern cluster. Continued monitoring of the avian disease is of critical importance to better understand and predict the epidemiological behaviour of the human cases.

Pathogenic influenza B virus in the ferret model establishes lower respiratory tract infection.

Influenza B viruses have become increasingly more prominent during influenza seasons. Influenza B infection is typically considered a mild disease and receives less attention than influenza A, but has been causing 20 to 50 % of the total influenza incidence in several regions around the world. Although there is increasing evidence of mid to lower respiratory tract diseases such as bronchitis and pneumonia in influenza B patients, little is known about the pathogenesis of recent influenza B viruses. Here we investigated the clinical and pathological profiles of infection with strains representing the two current co-circulating B lineages (B/Yamagata and B/Victoria) in the ferret model. Specifically, we studied two B/Victoria (B/Brisbane/60/2008 and B/Bolivia/1526/2010) and two B/Yamagata (B/Florida/04/2006 and B/Wisconsin/01/2010) strain infections in ferrets and observed strain-specific but not lineage-specific pathogenicity. We found B/Brisbane/60/2008 caused the most severe clinical illness and B/Brisbane/60/2008 and the B/Yamagata strains instigated pathology in the middle to lower respiratory tract. Importantly, B/Brisbane/60/2008 established efficient lower respiratory tract infection with high viral burden. Our phylogenetic analyses demonstrate profound reassortment among recent influenza B viruses, which indicates the genetic make-up of B/Brisbane/60/2008 differs from the other strains. This may explain the pathogenicity difference post-infection in ferrets.

Sequencing, annotation, and characterization of the influenza ferret infectome.

Ferrets have become an indispensable tool in the understanding of influenza virus virulence and pathogenesis. Furthermore, ferrets are the preferred preclinical model for influenza vaccine and therapeutic testing. Here we characterized the influenza infectome during the different stages of the infectious process in ferrets with and without prior specific immunity to influenza. RNA from lung tissue and lymph nodes from infected and naïve animals was subjected to next-generation sequencing, followed by de novo data assembly and annotation of the resulting sequences; this process generated a library comprising 13,202 ferret mRNAs. Gene expression profiles during pandemic H1N1 (pdmH1N1) influenza virus infection were analyzed by digital gene expression and solid support microarrays. As expected during primary infection, innate immune responses were triggered in the lung tissue; meanwhile, in the lymphoid tissue, genes encoding antigen presentation and maturation of effector cells of adaptive immunity increased dramatically. After 5 days postinfection, the innate immune gene expression was replaced by the adaptive immune response, which correlates with viral clearance. Reinfection with homologous pandemic influenza virus resulted in a diminished innate immune response, early adaptive immune gene regulation, and a reduction in clinical severity. The fully annotated ferret infectome will be a critical aid to the understanding of the molecular events that regulate disease severity and host-influenza virus interactions among seasonal, pandemic, and highly pathogenic avian influenzas.

Differential pathological and immune responses in newly weaned ferrets are associated with a mild clinical outcome of pandemic 2009 H1N1 infection.

Young children are typically considered a high-risk group for disease associated with influenza virus infection. Interestingly, recent clinical reports suggested that young children were the smallest group of cases with severe pandemic 2009 H1N1 (H1N1pdm) influenza virus infection. Here we established a newly weaned ferret model for the investigation of H1N1pdm infection in young age groups compared to adults. We found that young ferrets had a significantly milder fever and less weight loss than adult ferrets, which paralleled the mild clinical symptoms in the younger humans. Although there was no significant difference in viral clearance, disease severity was associated with pulmonary pathology, where newly weaned ferrets had an earlier pathology improvement. We examined the immune responses associated with protection of the young age group during H1N1pdm infection. We found that interferon and regulatory interleukin-10 responses were more robust in the lungs of young ferrets. In contrast, myeloperoxidase and major histocompatibility complex responses were persistently higher in the adult lungs; as well, the numbers of inflammation-prone granulocytes were highly elevated in the adult peripheral blood. Importantly, we observed that H1N1pdm infection triggered formation of lung structures that resembled inducible bronchus-associated lymphoid tissues (iBALTs) in young ferrets which were associated with high levels of homeostatic chemokines CCL19 and CXCL13, but these were not seen in the adult ferrets with severe disease. These results may be extrapolated to a model of the mild disease seen in human children. Furthermore, these mechanistic analyses provide significant new insight into the developing immune system and effective strategies for intervention and vaccination against respiratory viruses.

Seasonal H1N1 influenza virus infection induces cross-protective pandemic H1N1 virus immunity through a CD8-independent, B cell-dependent mechanism.

During the 2009 H1N1 influenza virus pandemic (pdmH1N1) outbreak, it was found that most individuals lacked antibodies against the new pdmH1N1 virus, and only the elderly showed anti-hemagglutinin (anti-HA) antibodies that were cross-reactive with the new strains. Different studies have demonstrated that prior contact with the virus can confer protection against strains with some degree of dissimilarity; however, this has not been sufficiently explored within the context of a pdmH1N1 virus infection. In this study, we have found that a first infection with the A/Brisbane/59/2007 virus strain confers heterologous protection in ferrets and mice against a subsequent pdmH1N1 (A/Mexico/4108/2009) virus infection through a cross-reactive but non-neutralizing antibody mechanism. Heterologous immunity is abrogated in B cell-deficient mice but maintained in CD8(-/-) and perforin-1(-/-) mice. We identified cross-reactive antibodies from A/Brisbane/59/2007 sera that recognize non-HA epitopes in pdmH1N1 virus. Passive serum transfer showed that cross-reactive sH1N1-induced antibodies conferred protection in naive recipient mice during pdmH1N1 virus challenge. The presence or absence of anti-HA antibodies, therefore, is not the sole indicator of the effectiveness of protective cross-reactive antibody immunity. Measurement of additional antibody repertoires targeting the non-HA antigens of influenza virus should be taken into consideration in assessing protection and immunization strategies. We propose that preexisting cross-protective non-HA antibody immunity may have had an overall protective effect during the 2009 pdmH1N1 outbreak, thereby reducing disease severity in human infections.

HAVOC: Small-scale histomic mapping of cancer biodiversity across large tissue distances using deep neural networks.

Intratumoral heterogeneity can wreak havoc on current precision medicine strategies because of challenges in sufficient sampling of geographically separated areas of biodiversity distributed across centimeter-scale tumor distances. To address this gap, we developed a deep learning pipeline that leverages histomorphologic fingerprints of tissue to create "Histomic Atlases of Variation Of Cancers" (HAVOC). Using a number of objective molecular readouts, we demonstrate that HAVOC can define regional cancer boundaries with distinct biology. Using larger tumor specimens, we show that HAVOC can map biodiversity even across multiple tissue sections. By guiding profiling of 19 partitions across six high-grade gliomas, HAVOC revealed that distinct differentiation states can often coexist and be regionally distributed within these tumors. Last, to highlight generalizability, we benchmark HAVOC on additional tumor types. Together, we establish HAVOC as a versatile tool to generate small-scale maps of tissue heterogeneity and guide regional deployment of molecular resources to relevant biodiverse niches.

CXCL10 contributes to p38-mediated apoptosis in primary T lymphocytes in vitro.

CXCL10 is part of the group of interferon-stimulated genes and it plays an important role during different viral infections by inducing cell activation, chemotaxis and lymphocyte priming toward the Th1 phenotype. In this study, we investigated in vitro the effects of CXCL10 in activated human primary T lymphocytes in terms of apoptosis or survival, and delineated the signaling pathways that are involved. CXCL10, in combination with IL-2 and/or IFNα, induces apoptosis in T lymphocytes. Moreover, CXCL10-induced activation of CXCR3 also triggers pro-survival signals that can be blocked by pertussis toxin. The analysis of the downstream signaling kinases shows that apoptosis is p38 MAPK-dependent and the pro-survival signals rely on the sustained activation of PI3K and the transient activation of Akt. On the other hand, the transient activation of p44/p42 ERK did not have an impact on T lymphocyte survival. We propose an immunological model in which CXCL10, together with other co-stimulating cytokines, participates in the activation of T lymphocytes, promotes survival and expansion of certain lymphocyte subsets, and induces chemotaxis toward the infected tissues. On the other hand, CXCL10 might contribute to the triggering of apoptosis in other subsets of T lymphocytes, including those lymphocytes that were transiently activated but later lacked the appropriate sets of specific co-stimulating signals to ensure their survival.

Heterogeneous virulence of pandemic 2009 influenza H1N1 virus in mice.

BACKGROUND: Understanding the pathogenesis of influenza infection is a key factor leading to the prevention and control of future outbreaks. Pandemic 2009 Influenza H1N1 infection, although frequently mild, led to a severe and fatal form of disease in certain cases that make its virulence nature debatable. Much effort has been made toward explaining the determinants of disease severity; however, no absolute reason has been established. RESULTS: This study presents the heterogeneous virulence of clinically similar strains of pandemic 2009 influenza virus in human alveolar adenocarcinoma cells and mice. The viruses were obtained from patients who were admitted in a local hospital in China with a similar course of infection and recovered. The A/Nanchang/8002/2009 and A/Nanchang/8011/2009 viruses showed efficient replication and high lethality in mice while infection with A/Nanchang/8008/2009 was not lethal with impaired viral replication, minimal pathology and modest proinflammatory activity in lungs. Sequence analysis displayed prominent differences between polymerase subunits (PB2 and PA) of viral genomes that might correlate with their different phenotypic behavior. CONCLUSIONS: The study confirms that biological heterogeneity, linked with the extent of viral replication, exists among pandemic H1N1 strains that may serve as a benchmark for future investigations on influenza pathogenesis.

Topographic mapping of the glioblastoma proteome reveals a triple-axis model of intra-tumoral heterogeneity.

Glioblastoma is an aggressive form of brain cancer with well-established patterns of intra-tumoral heterogeneity implicated in treatment resistance and progression. While regional and single cell transcriptomic variations of glioblastoma have been recently resolved, downstream phenotype-level proteomic programs have yet to be assigned across glioblastoma's hallmark histomorphologic niches. Here, we leverage mass spectrometry to spatially align abundance levels of 4,794 proteins to distinct histologic patterns across 20 patients and propose diverse molecular programs operational within these regional tumor compartments. Using machine learning, we overlay concordant transcriptional information, and define two distinct proteogenomic programs, MYC- and KRAS-axis hereon, that cooperate with hypoxia to produce a tri-dimensional model of intra-tumoral heterogeneity. Moreover, we highlight differential drug sensitivities and relative chemoresistance in glioblastoma cell lines with enhanced KRAS programs. Importantly, these pharmacological differences are less pronounced in transcriptional glioblastoma subgroups suggesting that this model may provide insights for targeting heterogeneity and overcoming therapy resistance.

Molecular characterization of in vivo adjuvant activity in ferrets vaccinated against influenza virus.

The 2009 H1N1 influenza pandemic has prompted a significant need for the development of efficient, single-dose, adjuvanted vaccines. Here we investigated the adjuvant potential of CpG oligodeoxynucleotide (ODN) when used with a human seasonal influenza virus vaccine in ferrets. We found that the CpG ODN-adjuvanted vaccine effectively increased antibody production and activated type I interferon (IFN) responses compared to vaccine alone. Based on these findings, pegylated IFN-alpha2b (PEG-IFN) was also evaluated as an adjuvant in comparison to CpG ODN and complete Freund's adjuvant (CFA). Our results showed that all three vaccines with adjuvant added prevented seasonal human A/Brisbane/59/2007 (H1N1) virus replication more effectively than did vaccine alone. Gene expression profiles indicated that, as well as upregulating IFN-stimulated genes (ISGs), CpG ODN enhanced B-cell activation and increased Toll-like receptor 4 (TLR4) and IFN regulatory factor 4 (IRF4) expression, whereas PEG-IFN augmented adaptive immunity by inducing major histocompatibility complex (MHC) transcription and Ras signaling. In contrast, the use of CFA as an adjuvant induced limited ISG expression but increased the transcription of MHC, cell adhesion molecules, and B-cell activation markers. Taken together, our results better characterize the specific molecular pathways leading to adjuvant activity in different adjuvant-mediated influenza virus vaccinations.

Downy mildew (Pl ( 8 ) and Pl ( 14 )) and rust (R ( Adv )) resistance genes reside in close proximity to tandemly duplicated clusters of non-TIR-like NBS-LRR-encoding genes on sunflower chromosomes 1 and 13.

Nucleotide binding site-leucine rich repeat (NBS-LRR) proteins are encoded by a ubiquitous gene family in sunflower and frequently harbor disease resistance genes. We investigated NBS-LRR-encoding resistance gene candidates (RGCs) flanking the downy mildew resistance genes Pl ( 8 ) and Pl ( 14 ) and the rust resistance gene R ( Adv ), which map on the NBS-LRR clusters of linkage groups 1 and 13 in sunflower genome. We shotgun sequenced bacterial artificial chromosome (BAC) clones proximal to Pl ( 8 ), Pl ( 14 ) , and R ( Adv ) and identified seven novel non-Toll/interleukin-1 receptor (TIR)-like NBS-LRR RGCs, which clustered with previously identified RGCs of linkage group 13 but were phylogenetically distant from the TIR- and non-TIR-NBS-LRR-encoding superfamilies of sunflower. Six of the seven predicted RGCs have intact open reading frames and reside in genomic segments with abundant transposable elements. The genomic localization and sequence similarity of the novel non-TIR-like predicted RGCs suggests that they originated from tandem duplications. RGCs in the proximity of Pl ( 8 ) and R ( Adv ) were likely introgressed from silverleaf sunflower genome, where the RGC cluster of linkage group 13 is duplicated in two independent chromosomes that have different architecture and level of recombination from the respective common sunflower chromosomes.

Raltitrexed-Modified Gold and Silver Nanoparticles for Targeted Cancer Therapy: Cytotoxicity Behavior In Vitro on A549 and HCT-116 Human Cancer Cells.

Two different raltitrexed gold and silver nanoparticles for the delivery of an antitumoral drug into cancer cells were synthesized and characterized. A cysteine linker was used for the covalent bonding of raltitrexed to the surface of nanoparticles. To evaluate the efficacy of the antifolate-derivative nanoparticles, their cytotoxicity was assayed in vitro with A549 human lung adenocarcinoma and HCT-116 colorectal carcinoma human cells. Modified nanoparticles are a biocompatible material, and administration of silver raltitrexed nanoparticles strongly inhibited the viability of the cancer cells; gold raltitrexed nanoparticles do not show any type of cytotoxic effect. The results suggest that silver raltitrexed nanoparticles could be a potential delivery system for certain cancer cells.

Unsupervised Resolution of Histomorphologic Heterogeneity in Renal Cell Carcinoma Using a Brain Tumor-Educated Neural Network.

PURPOSE: Applications of deep learning to histopathology have proven capable of expert-level performance, but approaches have largely focused on supervised classification tasks requiring context-specific training and deployment. More generalizable workflows that can be easily shared across subspecialties could help accelerate and broaden adoption. Here, we hypothesized that histology-optimized feature representations, generated by a convolutional neural network (CNN) during supervised learning, are transferable and can resolve meaningful differences in large-scale, discovery-type unsupervised analyses. METHODS: We used a CNN, previously trained to recognize brain tumor histomorphologies, to extract 512 feature representations from > 550 digital whole-slide images (WSIs) of renal cell carcinomas (RCCs) from The Cancer Genome Atlas and other previously unencountered tumors. We use these extracted feature vectors to conduct unsupervised image-set clustering and analyze the clinical and biologic relevance of the intra- and interpatient subgroups generated. RESULTS: Within individual WSIs, feature-based clustering could reliably segment tumor regions and other relevant histopathologic subpatterns (eg, adenosquamous and poorly differentiated regions). Across the larger RCC cohorts, clustering extracted features generated subgroups enriched for clinically relevant subtypes (eg, papillary RCC) and outcomes (eg, survival). Importantly, individual feature activation mapping highlighted salient subtype-specific patterns and features of malignancies (eg, nuclear grade, sarcomatous change) contributing to subgroupings. Moreover, some proposed clusters were enriched for recurring, human-based RCC-subtype misclassifications. CONCLUSION: Our data support that CNNs, pretrained on large histologic datasets, can extend learned representations to novel scenarios and resolve clinically relevant intra- and interpatient tissue-pattern differences without explicit instruction or additional optimization. Repositioning of existing histology-educated networks could provide scalable approaches for image classification, quality assurance, and discovery of unappreciated patterns and subgroups of disease.

High levels of proinflammatory cytokines, but not markers of tissue injury, in unaffected intestinal areas from patients with IBD.

Intestinal alterations in IBD are triggered and maintained by an overexpression of proinflammatory cytokines. Additionally, increased immune activation has been found in the adjacent intestinal areas without displaying any apparent histological alterations, however, the regulatory environment is not well established. Biopsy specimens from patients with ulcerative colitis (UC) and Crohn's disease (CD), from both affected and unaffected areas, and also from a group of colonic biopsies from healthy controls, were included in our study. Cytokines and markers of mucosal damage were analyzed by real-time PCR, and some of the results confirmed by western-blot and ELISA. Levels of IFNgamma, TNFalpha, IL-6, IL-15, IL-18, and IL-23 were increased (above healthy controls) in both affected and unaffected areas from IBD. IL-1beta, IL-6, IL-12, and IL-27 were higher in affected areas compared to unaffected ones in UC but not CD. In general, a correlation was observed between mRNA levels of these cytokines and both iNOS and Granzyme B. SOCS-2 and SOCS-3 were also increased in the affected areas. In conclusion, the unaffected areas from IBD show increased levels of a restricted set of cytokines that may exert immune activating roles in these areas without being able to trigger tissue damage.

Single nucleotide polymorphisms and linkage disequilibrium in sunflower.

Genetic diversity in modern sunflower (Helianthus annuus L.) cultivars (elite oilseed inbred lines) has been shaped by domestication and breeding bottlenecks and wild and exotic allele introgression(-)the former narrowing and the latter broadening genetic diversity. To assess single nucleotide polymorphism (SNP) frequencies, nucleotide diversity, and linkage disequilibrium (LD) in modern cultivars, alleles were resequenced from 81 genic loci distributed throughout the sunflower genome. DNA polymorphisms were abundant; 1078 SNPs (1/45.7 bp) and 178 insertions-deletions (INDELs) (1/277.0 bp) were identified in 49.4 kbp of DNA/genotype. SNPs were twofold more frequent in noncoding (1/32.1 bp) than coding (1/62.8 bp) sequences. Nucleotide diversity was only slightly lower in inbred lines ( = 0.0094) than wild populations ( = 0.0128). Mean haplotype diversity was 0.74. When extraploted across the genome ( approximately 3500 Mbp), sunflower was predicted to harbor at least 76.4 million common SNPs among modern cultivar alleles. LD decayed more slowly in inbred lines than wild populations (mean LD declined to 0.32 by 5.5 kbp in the former, the maximum physical distance surveyed), a difference attributed to domestication and breeding bottlenecks. SNP frequencies and LD decay are sufficient in modern sunflower cultivars for very high-density genetic mapping and high-resolution association mapping.

Host gene expression profiles in ferrets infected with genetically distinct henipavirus strains.

Henipavirus infection causes severe respiratory and neurological disease in humans that can be fatal. To characterize the pathogenic mechanisms of henipavirus infection in vivo, we performed experimental infections in ferrets followed by genome-wide gene expression analysis of lung and brain tissues. The Hendra, Nipah-Bangladesh, and Nipah-Malaysia strains caused severe respiratory and neurological disease with animals succumbing around 7 days post infection. Despite the presence of abundant viral shedding, animal-to-animal transmission did not occur. The host gene expression profiles of the lung tissue showed early activation of interferon responses and subsequent expression of inflammation-related genes that coincided with the clinical deterioration. Additionally, the lung tissue showed unchanged levels of lymphocyte markers and progressive downregulation of cell cycle genes and extracellular matrix components. Infection in the brain resulted in a limited breadth of the host responses, which is in accordance with the immunoprivileged status of this organ. Finally, we propose a model of the pathogenic mechanisms of henipavirus infection that integrates multiple components of the host responses.

Inhibition of influenza A virus infection by ginsenosides.

Influenza viruses cause mild to severe respiratory infections in humans. Due to efficient means of transmission, the viruses infect human population on a large scale. Apart from vaccines, antiviral drugs are used to control infection; neuraminidase inhibitors are thought to be the first choice of treatment, particularly for severe cases. Rapidly evolving and emerging influenza viruses with increased frequency of viral resistance to these drugs stress the need to explore novel antiviral compounds. In this study, we investigated antiviral activity of ginseng extract and ginsenosides, the ginseng-derived triterpene and saponin compounds, against 2009 pandemic H1N1 virus in vitro and in vivo. Our data showed that treatment of mice with ginsenosides protected the animals from lethal 2009 pandemic H1N1 infection and lowered viral titers in animal lungs. Mechanistic studies revealed that ginsenosides interact with viral hemagglutinin protein and prevent the attachment of virus with α 2-3' sialic acid receptors present on host cell surfaces. The interference in the viral attachment process subsequently minimizes viral entry into the cells and decreases the severity of the viral infection. We also describe that sugar moieties present in ginsenosides are indispensible for their attachment with viral HA protein. On the basis of our observations, we can say that ginsenosides are promising candidates for the development of antiviral drugs for influenza viruses.