Growth Kinetics Assay Research Articles

Antimicrobial activity of enacyloxin IIa and gladiolin against the urogenital pathogens Neisseria gonorrhoeae and Ureaplasma spp.

To determine the antimicrobial activity of enacyloxin IIa and gladiolin against Neisseria gonorrhoeae and Ureaplasma spp. The Burkholderia polyketide antibiotics enacyloxin IIa and gladiolin were tested against 14 N. gonorrhoeae and 10 Ureaplasma spp. isolates including multidrug-resistant N. gonorrhoeae isolates WHO V, WHO X and WHO Z as well as macrolide, tetracycline and ciprofloxacin-resistant ureaplasmas. Susceptibility testing of N. gonorrhoeae was carried out by agar dilution, whereas broth micro-dilution and growth kinetic assays were used for Ureaplasma spp. The MIC range for enacyloxin IIa and gladiolin against N. gonorrhoeae was 0·015-0·06mgl-1 and 1-2mgl-1 respectively. The presence of resistance to front line antibiotics had no effect on MIC values. The MIC range for enacyloxin IIa against Ureaplasma spp. was 4-32mgl-1 with a clear dose-dependent effect when observed using a growth kinetic assay. Gladiolin had no antimicrobial activity on Ureaplasma spp. at 32mgl-1 and limited impact on growth kinetics. Enacyloxin IIa and gladiolin antibiotics have antimicrobial activity against a range of antibiotic susceptible and resistant N. gonorrhoeae and Ureaplasma isolates. This study highlights the potential for a new class of antimicrobial against pathogens in which limited antibiotics are available. Development of these compounds warrants further investigation in the face of emerging extensively drug-resistant strains.

Resistance of a Halobacterium salinarum isolate from a solar saltern to cadmium, lead, nickel, zinc, and copper.

The current study focuses on the tolerance of a strain of Halobacterium salinarum isolated from Sfax solar saltern (Tunisia) towards cadmium (Cd), lead (Pb), nickel (Ni), zinc (Zn), and copper (Cu) by using agar dilution methods in complex and minimal media. The results showed the least inhibitory metals based on Minimum Inhibitory Concentrations (MICs) were lead (MIC = 4.5mM), cadmium (MIC = 4mM), and nickel (MIC = 2.5mM) in complex medium. The MICs of these metals were more inhibitory (MIC < 2mM) in the other tested media. The archaeal strain revealed a high sensitivity for copper and zinc, with MICs below 0.5mM for both metals. Growth kinetics in complex and minimal media showed the strain to be more sensitive to the metals in liquid media than in solid media. The growth kinetic assays indicated the presence of selected heavy metals resulted in a lower growth rate and lower total cell mass relative to the control. Despite that cadmium and lead are nonessential and have no nutrient value, they were the most tolerated metals by H. salinarum strain. In addition, pigment intensity in the strain was inhibited by the presence of the heavy metals relative to the control.

Growth kinetic and nitrogen source optimization for liquid culture fermentation of Metarhizium robertsii blastospores and bioefficacy against the corn leafhopper Dalbulus maidis.

The cosmopolitan entomopathogenic and root endophytic fungus Metarhizium robertsii has a versatile lifestyle and during liquid fermentation undergoes a dimorphic transformation from hyphae to conidia or microsclerotia, or from hyphae to blastospores. In all cases, these processes are mediated by environmental and nutritional cues. Blastospores could be used in spray applications to control arthropod pests above ground and may serve as an attractive alternative to the traditional solid-grown aerial conidial spores of Metarhizium spp. found in commercial products. Nitrogen is a vital nutrient in cell metabolism and growth; however, it is the expensive component in liquid cultures of entomopathogenic fungi. Our goals in this study were to optimize nitrogen sources and titers for maximum production of M. robertsii blastospores cultured in shake flasks at highly aerated conditions and to furtherdetermine theirvirulence against the corn leafhopper Dalbulus maidis, an important vector of serious pathogens in maize crops worldwide. Our fermentation studies revealed that the low-cost corn steep liquor (CSL) was the most suitable nitrogen source to improve blastospore growth in M. robertsii. The growth kinetic assays determined the optimal titer of 80gL-1 and a yield up to 4.7 × 108 cells mL-1 within 5days of cultivation (3days preculture and 2days culture), at a total cost of US$0.30L-1. Moreover, the blastospore growth kinetic was strongly dependent on glucose and nitrogen consumptions accompanied by a slight drop in the culture pH. Insect bioassays evidenced a high virulence of these blastospores, either as dried or fresh cells, to D. maidis adults fed on maize plants. Our findings provide insights into the nutritional requirements for optimal and cost-efficient production of M. robertsii blastospores and elucidate the potential of blastospores as an ecofriendly tool against the corn leafhopper.

Amino Acid Deletions in p6Gag Domain of HIV-1 CRF07_BC Ameliorate Galectin-3 Mediated Enhancement in Viral Budding.

HIV-1 CRF07_BC is a recombinant virus with amino acid (a.a.) deletions in p6Gag, which are overlapped with the Alix-binding domain. Galectin-3 (Gal3), a β-galactose binding lectin, has been reported to interact with Alix and regulate HIV-1 subtype B budding. This study aims to evaluate the role of Gal3 in HIV-1 CRF07_BC infection and the potential effect of a.a. deletions on Gal3-mediated regulation. A total of 38 HIV-1+ injecting drug users (IDUs) were enrolled in the study. Viral characterization and correlation of Gal3 were validated. CRF07_BC containing 7 a.a. deletions and wild-type in the p6Gag (CRF07_BC-7d and -wt) were isolated and infectious clones were generated. Viral growth kinetic and budding assays using Jurkat-CCR5/Jurkat-CCR5-Gal3 cells infected with CRF07_BC were performed. Results indicate that 69.4% (25/38) of the recruited patients were identified as CRF07_BC, and CRF07_BC-7d was predominant. Slow disease progression and significantly higher plasma Gal3 were noted in CRF07_BC patients (p < 0.01). Results revealed that CRF07_BC infection resulted in Gal3 expression, which was induced by Tat. Growth dynamic and budding assays indicated that Gal3 expression in Jurkat-CCR5 cells significantly enhanced CRF07_BC-wt replication and budding (p < 0.05), while the promoting effect was ameliorated in CRF07_BC-7d. Co-immunoprecipitation found that deletions in the p6Gag reduced Gal-3-mediated enhancement of the Alix–Gag interaction.

Propolis extract has bioactivity on the wall and cell membrane of Candida albicans

Ethnopharmacological relevanceThe use of natural products such as propolis extract (PE) is a promising alternative when topically administered to replace conventional antifungals, mostly due to its therapeutic applications, ease of access and low toxicity. However, despite being the subject of several mycology studies, they focus primarily on exploiting their antimicrobial activity, lacking information on the mechanisms of action of PE on Candida spp., characterizing its antifungal potential. Aim of the studyTo elucidate the bioactivity of PE on the cellular structure of Candida albicans. Materials and methodsA total of seven C. albicans clinical isolates plus a reference strain of C. albicans ATCC 90028 were used in this study. The PE was characterized and its effect on C. albicans was determined by susceptibility and growth kinetics assays; interference on C. albicans germination and filamentation; evaluation of the integrity of the C. albicans cell wall and membrane, as well as its mutagenic potential. ResultsThe PE presented strong inhibitory activity, which showed its greatest antifungal activity at 12 h with dose and time dependent fungistatic characteristics, effectively inhibiting and interfering on C. albicans filamentation. In addition, PE caused membrane and cell wall damage with intracellular content extravasation. Moreover, PE was not mutagenic. ConclusionsThe bioactivity of PE is mainly related to the loss of integrity membrane as well as the integrity of the cell wall and consequent increase in permeability, without mutagenic effects.

Facile One Pot Greener Synthesis of Sophorolipid Capped Gold Nanoparticles and its Antimicrobial Activity having Special Efficacy Against Gram Negative Vibrio cholerae

Microbes develop several strategies to survive in the adverse condition such as biofilm formation, attaining non-dividing state, altering drug target or drug, thereby increases the burden of drug dosage. To combat these issues, nanoparticles have shown an alternative approach for new treatment strategy but synthesis via chemical synthetic route limits their application in biomedical field. Here, green method for the synthesis of gold nanoparticles using sophorolipid (SL) is discussed that is characterized by various techniques. Initially, the antimicrobial activity was checked against metabolically active state of microbes; Gram-positive Staphylococcus aureus and Gram-negative Vibrio cholerae using XTT assay and growth kinetics assay. Results suggested higher efficacy of nanoparticles for Gram-negative, therefore further analyzed against Escherichia coli that confirmed its potency for the same. AuNPs-SL also signifies its efficiency at least metabolically active state; non dividing cells and biofilm of these microbes. Induced morphological changes were studied by SEM that revealed AuNPs-SL led to disruption of cell membrane and leakage of intracellular fluid to the surroundings. Inhibition of respiratory enzymes activity also plays a crucial role in bactericidal action as indicated by LDH assay. Synergy of AuNPs-SL with different antibiotics was also analyzed using checkerboard assay. These results suggested the possible use of AuNPs-SL as an antimicrobial therapy in the field of nanomedicine.

Micronized curcumin fabricated by supercritical CO2 to improve antibacterial activity against Pseudomonas aeruginosa

Curcumin (CM) is a natural polyphenolic compound with multiple biomedical functions. However, clinical applications face more challenges due to its low dissolution rate and poor bioavailability. Micronization is an effective strategy to overcome these drawbacks. Herein, CM nanoparticles (CM NPs, ∼300 nm) were fabricated using solution enhanced dispersion by supercritical CO2 (SEDS). The solubility of CM NPs was remarkably enhanced. Aim to study the effects of micronization on the biological functions of CM, we investigated the antibacterial activity of original CM and CM NPs upon Pseudomonas aeruginosa. In vitro, the minimal inhibitory concentrations (MIC) assay, solid-medium spot assay, growth kinetics assay and morphologic observation using atomic force microscopy (AFM) confirmed that the anti-P. aeruginosa activity of CM NPs was enhanced compared to original CM. Moreover, CM NPs also showed stronger inhibition for adhesion and biofilm formation of P. aeruginosa compared to original CM. Experiments on mice infected with P. aeruginosa showed that CM NPs have a better therapeutic effect than the original CM in vivo. In summary, CM NPs may be a novel and promising therapeutic candidate for bacterial infection.

Multi-Scale Dynamical Modelling of T-Cell Development from an Early Thymic Progenitor State to Lineage Commitment

Intrathymic development of committed pro-T-cells from multipotent hematopoietic precursors offers a unique opportunity to dissect the molecular circuitry establishing cell identity in response to environmental signals. This transition encompasses programmed shutoff of stem/progenitor genes, upregulation of T-cell specification genes, proliferation, and ultimately commitment. To explain these features in light of recently measured epigenetic effects and new experimental data, we developed a three-level dynamic model of commitment based upon regulation of the commitment-linked gene Bcl11b. The levels are: (1) a core gene regulatory network architecture determined by transcription factor (TF) perturbation data, (2) a stochastically controlled epigenetic gate, and (3) a single-cell proliferation model validated by experimental clonal growth and commitment kinetic assays. Using values from RNA-FISH measurements of genes encoding key TFs and bulk population dynamics, this single-cell model predicts state switching kinetics validated by measured clonal proliferation and commitment times. The resulting multi-scale model provides a novel approach and mechanistic framework for dissecting commitment dynamics.

Enhancing the production of scopoletin and quercetin 3-O-β-d-glucoside from cell suspension cultures of Tilia americana var. mexicana by modulating the copper and nitrate concentrations

The aerial tissues of Tilia americana var. mexicana produce compounds with anxiolytic activity, such as quercetin 3-O-β-d-glucoside and tiliroside, in addition to ones with anti-inflammatory properties, such as scopoletin. These three compounds were initially identified in callus cultures of apical buds. In the present study, suspension cultures from leaf explant callus were established; the accumulation of scopoletin and quercetin 3-O-β-d-glucoside in these cultures were found to be cell-growth-associated using cell growth and active compound-production kinetics assays. The effects of varying the nitrate and copper concentrations in Murashige and Skoog (MS, 27.4 mM total nitrates and 0.01 µM copper) medium on the growth of a suspension culture of T. americana cells and on the production of active compounds were tested by means of central composite design (CCD) generally used in the response surface methodology (RSM). Cell growth, measured as maximal biomass, improved when the total nitrate concentration decreased in the MS medium to 13.7 mM (p < 0.01) regardless of the copper concentration. As a phytoalexin, scopoletin accumulated rapidly in plants after pathogen infection, in the suspension cultures scopoletin yield was stimulated by increased copper concentration to 1.2 μM (p < 0.01). According to the C:N hypothesis, the carbon excess generated by nitrates reduced to 8.03 mM (p < 0.01) stimulated the production of quercetin 3-O-β-d-glucoside. Cell suspension of T. americana represents a potential biotechnological alternative for industrial exploitation in a stirred-tank bioreactor using a two-phase process: (1) the first step will be to grow the cell suspension, (2) the second stage will consist in handle the suspension culture towards the production of anxiolytic compounds or towards the production of anti-inflammatory compounds. As well as to evaluate another elicitors to stimulate tiliroside production in the T. americana suspension cultures.

Chondrogenic Progenitor Cells Exhibit Superiority Over Mesenchymal Stem Cells and Chondrocytes in Platelet-Rich Plasma Scaffold-Based Cartilage Regeneration

Background: Platelet-rich plasma (PRP) has been considered a promising tool for cartilage regeneration. However, increasing evidence has demonstrated the controversial effects of PRP on tissue regeneration, partially due to the unsatisfactory cell source. Chondrogenic progenitor cells (CPCs) have gained increasing attention as a potential cell source due to their self-renewal and multipotency, especially toward the chondrogenic lineage, and, thus, may be an appropriate alternative for cartilage engineering. Purpose: To compare the effects of PRP on CPC, mesenchymal stem cell (MSC), and chondrocyte proliferation, chondrogenesis, and cartilage regeneration. Study Design: Controlled laboratory study. Methods: Whole blood samples were obtained from 5 human donors to create PRPs (0, 1000 × 109, and 2000 × 109 platelets per liter). The proliferation and chondrogenesis of CPCs, bone marrow–derived MSCs (BMSCs), and chondrocytes were evaluated via growth kinetic and CCK-8 assays. Immunofluorescence, cytochemical staining, and gene expression analyses were performed to assess chondrogenic differentiation and cartilaginous matrix formation. The in vivo effects of CPCs, BMSCs, and chondrocytes on cartilage regeneration after PRP treatment were measured by use of histopathological, biochemical, and biomechanical techniques in a cartilage defect model involving mature male New Zealand White rabbits (critical size, 5 mm). Results: The CPCs possessed migration abilities and proliferative capacities superior to those of the chondrocytes, while exhibiting a chondrogenic predisposition stronger than that of the BMSCs. The growth kinetic, CCK-8, cytochemical staining, and biochemical analyses revealed that the CPCs simultaneously displayed a higher cell density than the chondrocytes and stronger chondrogenesis than the BMSCs after PRP stimulation. In addition, the in vivo study demonstrated that the PRP+CPC construct yielded better histological (International Cartilage Repair Society [ICRS] score, mean ± SEM, 1197.2 ± 163.2) and biomechanical (tensile modulus, 1.523 ± 0.194) results than the PRP+BMSC (701.1 ± 104.9, P < .05; 0.791 ± 0.151, P < .05) and PRP+chondrocyte (541.6 ± 98.3, P < .01; 0.587 ± 0.142, P < .01) constructs at 12 weeks after implantation. Conclusion: CPCs exhibit superiority over MSCs and chondrocytes in PRP scaffold-based cartilage regeneration, and PRP+CPC treatment may be a favorable strategy for cartilage repair. Clinical Relevance: These findings provide evidence highlighting the preferable role of CPCs as a cell source in PRP-mediated cartilage regeneration and may help researchers address the problem of unsatisfactory cell sources in cartilage engineering.

OMONITERING OF HEAVY METAL POLLUTION BY BIOLUMINESCENT BACTERIAL BIOSENSORS

Objectives: The current study focuses on the isolation of bioluminescent bacteria from the gut of Sea urchin and using its luminescence property as a potential biosensor for detecting chromium toxicity in water. Methodology: Bioluminescent bacteria, JAAKP J2 was isolated from the gut of Pseudoboletia indiana species of Sea urchin collected from Pondicherry university beach. The morphological, biochemical, enzyme profiling and molecular characterization through 16S sequencing were performed to identify the strain. Growth kinetic assays and spectrophotometric analysis under chromium stress condition were performed on bioluminescent strains for luminescence inhibition studies. The bioluminescent bacteria was then immobilized in nutrient agar cubesand used as a biosensor for the detection of hexavalent chromium concentration in water samples. Findings: The morphological, biochemical and molecular characterization revealed that the isolate JAAKP J2 was closely related to Vibrio campbellii. Industrially important enzymes like Protease, Lipase, Agarase, Cellulase, Xylanase and Gelatinase were also screened in this study. The developed biosensor using bioluminescence property of the isolate was able to detect the level of chromium toxicity in water samplesat concentration not more than 9 mg L-1 . Further we have elucidated a possible hypothetical pathway for reduction in luminescence property due to ROS (Reactive oxygen species) caused by hexavalent chromium toxicity in bacteria. Novelty: The study is first of its kind in isolation of bioluminescent strain from sea urchin and using it as a biosensor for heavy metal detection in water. Our study also throws lights on futuristic approach of detecting other heavy metals like lead, zinc, cadmium, mercury in water samples by bacterial luminescence. Keywords: Bioluminescent Bacteria, Biosensor, Chromium Toxicity, Enzyme Profiling, Sea Urchin

Isolation and biological characteristics of sheep amniotic epithelial cells.

Amniotic epithelial cells (AECs), isolated from placenta, have epithelial cells and stem cells characteristics. Most of the previous studies focused on the biological characteristics of human amniotic epithelial cells, which demonstrated amniotic epithelial cells not only had low immunogenicity and potent potential to differentiate into three germ layers, but also could secrete various immunomodulatory factors. However, compared to study on human amniotic epithelial cells, few studies have been done on other animals. In this study, sheep amniotic epithelial cells were successfully isolated and their surface makers were accessed by immunofluorescence assay, and found that AECs were expressed Oct4 and Sox2, which were necessary for maintaining the undifferentiated state of pluripotent stem cells. Based on cloning efficiency and growth kinetics assay, AECs were found to possess self-renewal capacity and the growth curve was S-shaped. In addition, AECs could be induced into adipocytes, osteoblasts and chondrocytes in vitro, showing they had multi-differential ability. Reverse transcription-polymerase chain reaction results showed that AECs expressed CD29, CD44, CD90 and CK19, and didn't expressed CD34, CD45 and the telomerase gene (TERT). Little change in chromosome number was observed in AEC cultures for up to at least the first ten passages. In summary, this study results revealed that sheep AECs possessed more advantages for cell therapy and might play a key role in cell therapy and regenerative medicine in the future.

Evolutionary evidence for multi-host transmission of cetacean morbillivirus

Cetacean morbillivirus (CeMV) has emerged as the pathogen that poses the greatest risk of triggering epizootics in cetacean populations worldwide, and has a high propensity for interspecies transmission, including sporadic infection of seals. In this study, we investigated the evolutionary history of CeMV by deep sequencing wild-type viruses from tissue samples representing cetacean species with different spatiotemporal origins. Bayesian phylogeographic analysis generated an estimated evolutionary rate of 2.34 × 10−4 nucleotide substitutions/site/year and showed that CeMV evolutionary dynamics are neither host-restricted nor location-restricted. Moreover, the dolphin morbillivirus strain of CeMV has undergone purifying selection without evidence of species-specific mutations. Cell-to-cell fusion and growth kinetics assays demonstrated that CeMV can use both dolphin and seal CD150 as a cellular receptor. Thus, it appears that CeMV can readily spread among multiple cetacean populations and may pose an additional spillover risk to seals.

The atrial appendage as a suitable source to generate cardiac-derived adherent proliferating cells for regenerative cell-based therapies.

Cardiac-derived adherent proliferating (CardAP) cells obtained from endomyocardial biopsies (EMBs) with known anti-fibrotic and pro-angiogenic properties are good candidates for the autologous therapy of end-stage cardiac diseases such as dilated cardiomyopathy. However, due to the limited number of CardAP cells that can be obtained from EMBs, our aim is to isolate cells with similar properties from other regions of the heart with comparable tissue architecture. Here, we introduce the atrial appendage as a candidate region. Atrial appendage-derived cells were sorted with CD90 microbeads to obtain a CD90low cell population, which were subsequently analysed for their surface marker and gene expression profiles via flow cytometry and micro array analysis. Enzyme-linked immunosorbent assays for vascular endothelial growth factor and interleukin-8 as well as tube formation assays were performed to investigate pro-angiogenic properties. Furthermore, growth kinetic assays were performed to estimate the cell numbers needed for cell-based products. Microarray analysis revealed the expression of numerous pro-angiogenic genes and strong similarities to CardAP cells with which they also share expression levels of defined surface antigens, that is, CD29+ , CD44+ , CD45- , CD73+ , CD90low , CD105+ , and CD166+ . High secretion levels of vascular endothelial growth factor and interleukin-8 as well as improved properties of vascular structures in vitro could be detected. Based on growth parameters, cell dosages for the treatment of more than 250 patients are possible using one appendage. These results lead to the conclusion that isolating cells with regenerative characteristics from atrial appendages is feasible and permits further investigations towards allogenic cell-based therapies.

Allogenic human serum, a clinical grade serum supplement for promoting human periodontal ligament stem cell expansion.

Exposing human periodontal ligament stem cells (hPDLSCs) to animal proteins during cell expansion would compromise quality and safety of the hPDLSCs for clinical applications. The current study aimed to evaluate the replacement of animal-based serum by human serum for the expansion of hPDLSCs. hPDLSCs were cultured in culture media supplemented with four types of serums: Group A: fetal bovine serum (FBS); Group B: allogeneic human male AB serum (HS); Group C: in-house autologous (Auto-HS); and Group D: in-house allogeneic human serums (Allo-HS). Exhibitions of mesenchymal stem cell characteristics of hPDLSCs were examined. Then, growth and osteogenic (OS) differentiation potential of hPDLSCs in FBS and HS at passages 5 and 15 were compared to investigate the effects of serum supplements on growth and expansion stability of the expanded hPDLSCs. After that, growth and OS differentiation of hPDLSCs in Auto- and Allo-HS were investigated. Flow cytometrical analyses, functional differentiations, cell growth kinetic, cytogenetic analysis, alkaline phosphatase and calcium content assays, and oil red O and von Kossa staining were performed. Results showed that at passage 5, HS promoted growth and OS differentiation of hPDLSCs and extensive cell expansion, decreased growth and differentiation potential of the expanded hPDLSCs, particularly in HS. Growth and OS differentiation of hPDLSCs in Auto-HS and Allo-HS were not different. In summary, allogeneic human serum could be a replacement to FBS for hPDLSC expansion. In vitro cell expansion of hPDLSCs should be minimal to ensure optimal cell quality. Copyright © 2016 John Wiley & Sons, Ltd.

Double Plant Homeodomain Fingers 2 (DPF2) Promotes the Immune Escape of Influenza Virus by Suppressing Beta Interferon Production.

The high mutation rates of the influenza virus genome facilitate the generation of viral escape mutants, rendering vaccines and drugs against influenza virus-encoded targets potentially ineffective. Therefore, we identified host cell determinants dispensable for the host but crucial for virus replication, with the goal of preventing viral escape and finding effective antivirals. To identify these host factors, we screened 2,732 human genes using RNA interference and focused on one of the identified host factors, the double plant homeodomain fingers 2 (DPF2/REQ) gene, for this study. We found that knockdown of DPF2 in cells infected with influenza virus resulted in decreased expression of viral proteins and RNA. Furthermore, production of progeny virus was reduced by two logs in the multiple-cycle growth kinetics assay. We also found that DPF2 was involved in the replication of seasonal influenza A and B viruses. Because DPF2 plays a crucial role in the noncanonical NF-κB pathway, which negatively regulates type I interferon (IFN) induction, we examined the relationship between DPF2 and IFN responses during viral infection. The results showed that knockdown of DPF2 resulted in increased expression of IFN-β and induced phosphorylation of STAT1 in infected cells. In addition, high levels of several cytokines/chemokines (interleukin-8 [IL-8], IP-10, and IL-6) and antiviral proteins (MxA and ISG56) were produced by DPF2 knockdown cells. In conclusion, we identified a novel host factor, DPF2, that is required for influenza virus to evade the host immune response and that may serve as a potential antiviral target.IMPORTANCE Influenza virus is responsible for seasonal epidemics and occasional pandemics and is an ongoing threat to public health worldwide. Influenza virus relies heavily on cellular factors to complete its life cycle. Here we identified a novel host factor, DPF2, which is involved in influenza virus infection. Our results showed that DPF2 plays a crucial role in the replication and propagation of influenza virus. DPF2 functions in the noncanonical NF-κB pathway, which negatively regulates type I IFN induction. Thus, we investigated the relationship between the IFN response and DPF2 in influenza virus infection. Upon influenza virus infection, DPF2 dysregulated IFN-β induction and expression of cytokines/chemokines and antiviral proteins. This study provides evidence that influenza virus utilizes DPF2 to escape host innate immunity.

Biomimetic synthesis of AgNPs from Penicillium chrysogenum strain FGCC/BLS1 by optimising physico-cultural conditions and assessment of their antimicrobial potential.

Biomimetic synthesis of metal nanoparticles (NPs) is safe and eco-friendly; therefore, find diverse applications. Considering this, the soil fungi Penicillium chrysogenum strain Fungal germplasm collection centre/ BLS1 was isolated, characterized and explored to synthesize extracellular silver NPs (AgNPs) under optimised conditions. The synthesis of AgNPs was investigated using ultraviolet (UV)-visible spectroscopy, Fourier-transform infra-red spectroscopy (FTIR), transmission electron microscope (TEM) and dynamic light scattering (DLS) analysis. Process optimisation exhibited AgNPs synthesis within 8 h using 2 mM AgNO3 at pH 11 and temperature 70°C. TEM analysis revealed polydispersed ellipsoidal shaped AgNPs with average particle size 96.8 nm as measured by DLS. AgNPs showed negative zeta potential that confers surface stability in solution. FTIR spectra confirmed the presence of protein bound to AgNPs. Antibacterial activity against Escherichia coli, Klebsiella pneumoniae and Staphylococcus aureus by the AgNPs (100 ppm) was demonstrated by counting colony forming unit , disc diffusion, and growth kinetics assay. Additionally radial assay revealed antifungal activity of AgNPs (100 ppm) against phytopathogenic fungi Sclerotinia sclerotiorum Microbial type culture collection 8785. Furthermore, AgNPs (100 ppm) did not show any cytotoxic effects on human Red blood cells. Therefore, this novel fungal strain can be utilised for biofabrication of AgNPs under optimised conditions and have shown strong antimicrobial property.

Genomic regions, cellular components and gene regulatory basis underlying pod length variations in cowpea (V.unguiculata L. Walp).

SummaryCowpea (V. unguiculata L. Walp) is a climate resilient legume crop important for food security. Cultivated cowpea (V. unguiculata L) generally comprises the bushy, short‐podded grain cowpea dominant in Africa and the climbing, long‐podded vegetable cowpea popular in Asia. How selection has contributed to the diversification of the two types of cowpea remains largely unknown. In the current study, a novel genotyping assay for over 50 000 SNPs was employed to delineate genomic regions governing pod length. Major, minor and epistatic QTLs were identified through QTL mapping. Seventy‐two SNPs associated with pod length were detected by genome‐wide association studies (GWAS). Population stratification analysis revealed subdivision among a cowpea germplasm collection consisting of 299 accessions, which is consistent with pod length groups. Genomic scan for selective signals suggested that domestication of vegetable cowpea was accompanied by selection of multiple traits including pod length, while the further improvement process was featured by selection of pod length primarily. Pod growth kinetics assay demonstrated that more durable cell proliferation rather than cell elongation or enlargement was the main reason for longer pods. Transcriptomic analysis suggested the involvement of sugar, gibberellin and nutritional signalling in regulation of pod length. This study establishes the basis for map‐based cloning of pod length genes in cowpea and for marker‐assisted selection of this trait in breeding programmes.

Two potential recombinant rabies vaccines expressing canine parvovirus virion protein 2 induce immunogenicity to canine parvovirus and rabies virus

Both rabies virus (RABV) and canine parvovirus (CPV) cause lethal diseases in dogs. In this study, both high egg passage Flury (HEP-Flury) strains of RABV and recombinant RABV carrying double RABV glycoprotein (G) gene were used to express the CPV virion protein 2 (VP2) gene, and were designated rHEP-VP2 and, rHEP-dG-VP2 respectively. The two recombinant RABVs maintained optimal virus titration according to their viral growth kinetics assay compared with the parental strain HEP-Flury. Western blotting indicated that G protein and VP2 were expressed in vitro. The expression of VP2 in Crandell feline kidney cells post-infection by rHEP-VP2 and rHEP-dG-VP2 was confirmed by indirect immunofluorescence assay with antibody against VP2. Immunogenicity of recombinant rabies viruses was tested in Kunming mice. Both rHEP-VP2 and rHEP-dG-VP2 induced high levels of rabies antibody compared with HEP-Flury. Mice immunized with rHEP-VP2 and rHEP-dG-VP2 both had a high level of antibodies against VP2, which can protect against CPV infection. A challenge experiment indicated that more than 80% mice immunized with recombinant RABVs survived after infection of challenge virus standard 24 (CVS-24). Together, this study showed that recombinant RABVs expressing VP2 induced protective immune responses to RABV and CPV. Therefore, rHEP-VP2 and rHEP-dG-VP2 might be potential combined vaccines for RABV and CPV.

Process optimization for green synthesis of silver nanoparticles by Sclerotinia sclerotiorum MTCC 8785 and evaluation of its antibacterial properties.

BackgroundEco-friendly synthesis of nanoparticles is viewed as an alternative to the chemical method and initiated the use of microorganisms for synthesis. The present study has been designed to utilize plant pathogenic fungi Sclerotinia sclerotiorum MTCC 8785 strain for synthesis and optimization of silver nanoparticles (AgNPs) production as well as evaluation of antibacterial properties. The AgNPs were synthesized by reduction of aqueous silver nitrate (AgNO3) solution after incubation of 3–5 days at room temperature. The AgNPs were further characterized using UV–visible spectroscopy, Fourier transform infrared spectroscopy (FTIR) and transmission electron microscopy (TEM). Reaction parameters including media, fungal biomass, AgNO3 concentration, pH and temperature were further optimized for rapid AgNPs production. The antibacterial efficacy of AgNPs was evaluated against Escherichia coli ATCC 25922 and Staphylococcus aureus ATCC 25923 by disc diffusion and growth kinetics assay at the concentration determined by the minimum inhibitory concentration (MIC).ResultsAgNPs synthesis was initially marked by the change in colour from pale white to brown and was confirmed by UV–Vis spectroscopy. Optimization studies showed that potato dextrose broth (PDB) media, 10 g of biomass, addition of 2 mM AgNO3, pH 11 and 80 °C temperature resulted in enhanced AgNPs synthesis through extracellular route. TEM data revealed spherical shape AgNPs with size in the range of 10 nm. Presence of proteins capped to AgNPs was confirmed by FTIR. AgNPs showed antibacterial activity against E. coli and S. aureus at 100 ppm concentration, corresponding MIC value.ConclusionS. sclerotiorum MTCC 8785 mediated AgNPs was synthesized rapidly under optimized conditions, which showed antibacterial activity.Electronic supplementary materialThe online version of this article (doi:10.1186/s40064-016-2558-x) contains supplementary material, which is available to authorized users.