Electromagnetic Irradiation Evokes Physiological and Molecular Alterations in Rice.

Electromagnetic energy is the "backbone" of wireless communication systems, and its progressive use is considered to have a low but measurable impact on a wide range of biological systems. Even though a growing amount of data has reported electromagnetic energy absorption in humans along with subsequent biological effects, the consequences of electromagnetic energy absorption on plants have been insufficiently addressed. The higher surface to volume ratio along with the enormous water-ion concentrations makes the plant an ideal model to interact with non-ionizing electromagnetic radiation. In this study, controlled and periodic electromagnetic exposure of 1837.50 MHz, 2.75 W/m2 for 6 h a day on a popular rice variety (var. Satabdi) reduced the seed germination rate. The same dose of periodic electromagnetic exposure upregulated phytochrome B and phytochrome C gene transcripts in 12-day-old seedlings, whereas, in 32-day-old plants, the dose upregulated calmodulin and phytochrome C while the bZIP1 gene showed repression. However, the transcript abundance of bZIP1, phytochrome B, and phytochrome C genes was enhanced even in 12-day-old Satabdi seedlings following instantaneous short-duration (2 h 30 min) controlled electromagnetic exposure to 1837.50 MHz, 2.75 W/m2 . The reported responses in rice were observed below the international electromagnetic regulatory limits. Thus, rice plants perceived electromagnetic energy emitted by the wireless communication system as abiotic stress as per its response by upregulation or repression of known stress-sensing genes. Bioelectromagnetics. © 2020 Bioelectromagnetics Society.

One-time Electromagnetic Irradiation Modifies Stress-sensitive Gene Expressions in Rice Plant.

Electromagnetic energy is utilized over multiple frequency bands to provide seamless wireless communication services. Plants can well perceive electromagnetic energy present in open environment due to reasonably high permittivity and electrical conductivity of constituent tissues. Moreover, higher surface-to-volume ratio of plant structure facilitates increased interaction with the incident electromagnetic waves. To date, a few well-designed studies have been conducted inside controlled electromagnetic reverberation chambers to investigate either short duration-low amplitude or long duration-periodic electromagnetic irradiation-induced molecular responses in plants. However, as far as is known, studies investigating molecular responses particularly at the mid-vegetative stage in plants following one-time (hours-long) electromagnetic irradiation have not been reported earlier. Hence, the present study aimed at investigating molecular responses in 40-day-old Swarnaprabha rice plants following one-time 1837.50 MHz, 2.75 mW/m2 electromagnetic irradiation of 2 h 30 min duration. Controlled electromagnetic irradiation inside a simple reverberation chamber was ensured to achieve pure electromagnetic environment at 1837.50 MHz with deterministic electromagnetic power density at selected position. Swarnaprabha rice plant was chosen for this investigation since the rice variety is widely cultivated and consumed in the Indian subcontinent. Subsequent alterations in some selected stress-sensitive gene expressions were assayed using real-time quantitative polymerase chain reaction technique-significant upregulation in calmodulin and phytochrome B gene expressions were noted. This investigation was purposefully focused on subsequent molecular responses immediately following electromagnetic irradiation so that the possible effects of secondary stimulations could be avoided. Observed molecular responses strongly suggested that plants perceive 1837.50 MHz, 2.75 mW/m2 electromagnetic irradiation similar to other injurious stimuli. © 2021 Bioelectromagnetics Society.

Translation of Mycobacterium Survival Strategy to Develop a Lipo-peptide based Fusion Inhibitor*.

The entry of enveloped virus requires the fusion of viral and host cell membranes. An effective fusion inhibitor aiming at impeding such membrane fusion may emerge as a broad-spectrum antiviral agent against a wide range of viral infections. Mycobacterium survives inside the phagosome by inhibiting phagosome-lysosome fusion with the help of a coat protein coronin 1. Structural analysis of coronin 1 and other WD40-repeat protein suggest that the trp-asp (WD) sequence is placed at distorted ß-meander motif (more exposed) in coronin 1. The unique structural feature of coronin 1 was explored to identify a simple lipo-peptide sequence (myr-WD), which effectively inhibits membrane fusion by modulating the interfacial order, water penetration, and surface potential. The mycobacterium inspired lipo-dipeptide was successfully tested to combat type 1 influenza virus (H1N1) and murine coronavirus infections as a potential broad-spectrum antiviral agent.

Aqua-friendly organometallic Ir-Pt complexes: pH-responsive AIPE-guided imaging of bacterial cells.

In this work, the aggregation-induced photoluminescence emission (AIPE) of three water-soluble heterobimetallic Ir-Pt complexes was reported with insight into their photophysical and electrochemical properties and imaging of bacterial cells. An alkyne appended Schiff's base L, bridges bis-cyclometalated iridium(III) and platinum(II) terpyridine centre. The Schiff's base (N-N fragment) serves as the ancillary ligand to the iridium(III) centre, while the alkynyl end is coordinated to platinum(II). The pH and ionic strength influence the aggregation kinetics of the alkynylplatinum(II) fragment, leading to metal-metal and π-π interactions with the emergence of a triplet metal-metal-to-ligand charge transfer (3MMLCT) emission. The excellent reversibility and photostability of aggregation-induced emission (AIE) of these aqua-friendly complexes were tested for their ability to sense and selectively image E. coli cells at various pH values.

Heterogeneity and Compositional Diversities of Campylobacter jejuni Outer Membrane Vesicles (OMVs) Drive Multiple Cellular Uptake Processes.

Naturally secreted outer membrane vesicles (OMVs) from gut microbes carry diverse cargo, including proteins, nucleic acids, toxins, and many unidentified secretory factors. Bacterial OMVs can shuttle molecules across different cell types as a generalized secretion system, facilitating bacterial pathogenicity and self-survival. Numerous mucosal pathogens, including Campylobacter jejuni (C. jejuni), share a mechanism of harmonized secretion of major virulence factors. Intriguingly, as a common gut pathogen, C. jejuni lacks some classical virulence-associated secretion systems; alternatively, it often employs nanosized lipid-bound OMVs as an intensive strategy to deliver toxins, including secretory proteins, into the target cells. To better understand how the biophysical and compositional attributes of natural OMVs of C. jejuni regulate their cellular interactions to induce a biologically relevant host response, we conducted an in-depth morphological and compositional analysis of naturally secreted OMVs of C. jejuni. Next, we focused on understanding the mechanism of host cell-specific OMVs uptake from the extracellular milieu. We showed that intracellular perfusion of OMVs is mediated by cytosolic as well as multiple endocytic uptake processes due to the heterogenic nature, abundance of surface proteins, and membrane phospholipids acquired from the source bacteria. Furthermore, we used human and avian cells as two different host targets to provide evidence of target cell-specific preferential uptake of OMVs. Together, the present study provides insight into the unique functionality of natural OMVs of C. jejuni at the cellular interface, upholding their potential for multimodal use as prophylactic and therapeutic carriers.

Gut Microbe-Derived Outer Membrane Vesicles: A Potential Platform to Control Cecal Load of Campylobacter jejuni.

Acute diarrheal illness and gastroenteritis caused by Campylobacter jejuni infection remain significant public health risks in developing countries with substantial mortality and morbidity in humans, particularly in children under the age of five. Genetic diversities among Campylobacter jejuni and limited understanding of immunological correlations of host protection remain primary impediments for developing an effective measure to controlCampylobacter infection. Moreover, the lack of a reliable in vivo model to mimic natural infection against Campylobacter jejuni has substantially delayed the vaccine-development process. Given the role of bacterial outer membrane associated proteins in intestinal adherence and invasion as well as modulating dynamic interplay between host and pathogens, bacterial outer-membrane vesicles have emerged as a potential vaccine target against a number of gut pathogens, including Campylobacter jejuni. Here, we describe a mucosal vaccine strategy using chitosan-coated outer-membrane vesicles to induce specific immune responses against Campylobacter jejuni in mice. To overcome the challenges of mucosal delivery of outer membrane vesicles in terms of exposure to variable pH and risk of enzymatic degradation, we preferentially used chitosan as a nontoxic, mucoadhesive polymer. We show that intragastric delivery of chitosan-coated outer-membrane vesicles imparts significant immune protection against Campylobacter jejuni with high level local and systemic antibody production. Further, immunization with the outer membrane vesicles resulted in potent cellular responses with an increased CD4+ and CD8+ T cell population. Moreover, significant upregulation of IFN-γ and IL-6 gene expression suggests that mucosal delivery of outer membrane vesicles promotes a Th1/Th2 mixed-type immune response. Together, as an acellular and nonreplicating canonical end product of bacterial secretion, mucosal delivery of outer membrane vesicles may represent a promising platform for developing an effective vaccine againstCampylobacter jejuni.

Mucosal delivery of live Lactococcus lactis expressing functionally active JlpA antigen induces potent local immune response and prevent enteric colonization of Campylobacter jejuni in chickens.

Successful colonization of the mucosal epithelial cells is the key early step for Campylobacter jejuni (C. jejuni) pathogenesis in humans. A set of Surface Exposed Colonization Proteins (SECPs) are known to take leading role in bacterial adhesion and subsequent host pathogenesis. Among the major SECPs, the constitutively expressed C. jejuni surface lipoprotein Jejuni lipoprotein A (JlpA), interacts with intestinal heat shock protein 90α (Hsp90α) and contributes in disease progression by triggering pro-inflammatory responses via activation of NF-κB and p38 MAP kinase pathways. In addition to its ability to express on the surface, high sequence conservation of JlpA protein among different Campylobacter spp make it a suitable vaccine target against C. jejuni. Given that chickens are the primary source for C. jejuni infection in humans and persistent cecal colonization significantly contribute in pathogen transmission, we explicitly used chickens as a model to test the immune-protective efficacy of JlpA protein. Taking into account that gastro-intestinal tract is the major site for C. jejuni colonization, we chose to use mucosal (intragastric) route as mode for JlpA antigen delivery. To deliver JlpA via mucosal route, we engineered a food grade Lactic acid producing bacteria, Lactococcus lactis (L. lactis) to express functionally active JlpA protein in the surface. Further, we demonstrated its ability to substantially improve the antigen specific local immune responses in the intestine along with significant immune-protection against enteric colonization of C. jejuni in chickens.

Vaccination with CpG-adjuvanted avian influenza virosomes promotes antiviral immune responses and reduces virus shedding in chickens.

The use of virosomes as a vaccine platform has proven successful against several viruses. Here we examined the protective efficacy of a virosome-based vaccine consisting of avian influenza virus (AIV) A/Duck/Czech/56/H4N6 in chickens against a homologous AIV challenge. Virosomes adjuvanted with CpG-ODN or recombinant chicken interferon (IFN)-γ significantly reduced virus shedding after virus challenge. Furthermore, immunization with virosomes adjuvanted with CpG-ODN increased hemagglutination inhibition (HI) and virus-specific neutralizing serum antibodies, as well as virus-specific serum IgG and mucosal IgA responses. We also found a significant increase in the expression of type I and II interferon genes in the protected birds following virus challenge. In summary, this study demonstrated the ability of virosomes adjuvanted with CpG-ODN to reduce AIV shedding, and elicit virus-specific protective antibody responses in vaccinated birds.

Prophylactic treatment with Toll-like receptor ligands enhances host immunity to avian influenza virus in chickens.

Avian influenza viruses (AIV) pose a threat towards the health of both poultry and humans. To interrupt the transmission of the virus, novel prophylactic strategies must be considered which may reduce the shedding of AIV. One potential is the prophylactic use of Toll-like receptor (TLR) ligands. Many cells of the immune system express TLRs, and cellular responses to TLR stimulation include activation and the production of cytokines. TLR ligands have been employed as prophylactic treatments to enhance host resistance to pathogens both in mammals and chickens. Therefore, the present study was conducted to determine whether TLR ligands may be used prophylactically in chickens to enhance host immunity to AIV. Chickens received intramuscular injections of either low or high doses of the TLR ligands poly I:C, lipopolysaccharide (LPS) and CpG ODN. Twenty-four hours post-treatment, chickens were infected with the low pathogenic avian influenza virus H4N6, and both oropharyngeal and cloacal virus shedding were assessed on days 4 and 7 post-infection. To identify potential correlates of immunity, spleen and lungs were collected on days 2, 4 and 7 post-infection for RNA extraction. The results suggested that all of the TLR ligand treatments induced a significant reduction in virus shedding, with the TLR3 ligand poly I:C conferring the greatest AIV immunity compared to control birds, followed by CpG ODN and LPS. Furthermore, transcriptional analysis of gene expression in the spleen and lungs suggest IFN-α and IL-8 as correlates of immunity conferred by poly I:C, and IFN-γ for CpG ODN and LPS. In conclusion, TLR ligands, have the ability to enhance host immunity against AIV, and future studies should consider exploring the combinatory effects of poly I:C and CpG ODN prophylaxis in conjunction with AIV vaccination.

Interferon-γ influences immunity elicited by vaccines against very virulent Marek's disease virus.

Vaccination of chickens with herpesvirus of turkey (HVT) confers only partial protection against challenge with a very virulent Marek's disease virus (MDV). Here, we evaluated the ability of recombinant chicken interferon-gamma (rChIFN-γ) to enhance protective efficacy of HVT against the very virulent MDV strain, RB1B. The bioactivity of IFN-γ expressed by a plasmid expression vector was confirmed by its ability to stimulate a chicken macrophage cell line (HD11) to produce nitric oxide (NO) in vitro. The administration of HVT with 5µg of pcDNA:chIFN-γ plasmid reduced the incidence of tumor development significantly when compared to vaccinated birds (77.7% in the HVT+empty vector group and 80% in HVT group versus 33.3% in the HVT+chIFN-γ group) and significantly increased IFN-γ expression in the splenocytes of the protected group, suggesting that rChIFN-γ increases the potency of HVT against MDV. Further analysis demonstrated that the protected birds that received HVT vaccine and/or plasmid had lower MDV genome load and lower amounts of transcripts for meq and vIL-8 than in the birds without lesions. Similarly, lower expression of IL-10, IL-18 and IL-6 was observed in the chickens without lesions compared to the chickens that had lesions, suggesting an inverse association between up-regulation of these cytokines and vaccine-induced immunity. In conclusion, IFN-γ can positively influence immunity conferred by HVT vaccination against challenge with a very virulent Marek's disease virus (vvMDV) in chickens.

Assessment of bioactivity of a recombinant chicken interferon-gamma expressed using a baculovirus expression system.

The full-length coding sequence of chicken interferon-γ (ChIFN-γ) was cloned into a baculovirus nonfusion vector, pFastBacDual, and expressed in Sf21 insect cells. Recombinant ChIFN-γ (rChIFN-γ) protein was found to be expressed both intracellularly as well as in the culture supernatants. The affinity-purified rChIFN-γ contained 14, 17, and 28 kDa proteins, possibly representing both glycosylated and nonglycosylated protein forms of ChIFN-γ. The bioactivity of rChIFN-γ was confirmed in vitro by production of nitric oxide in a chicken macrophage cell line (HD11) and antiviral activity against vesicular stomatitis virus in primary chicken embryonic fibroblast cells. Further, HD11 cells stimulated with rChIFN-γ showed significant upregulation of inducible nitric oxide synthases, IFN-γ, interleukin-1ß, interleukin-12p35, signal transducers and activators of transcription 1, class II, major histocompatibility complex, transactivator, and major histocompatibility complex II-ß chain (BL-B) transcripts. In conclusion, the present study provides information on the ability of functionally active rChIFN-γ expressed in a baculovirus system in inducing significant transcriptional upregulation of various immune system-related genes, including those that encode cytokines, antigen-presenting molecules, and transcription factors involved in the major histocompatibility complex and IFN-signaling pathway.

Oral treatment of chickens with lactobacilli influences elicitation of immune responses.

Commensal microbes in the intestine are in constant interaction with host cells and play a role in shaping the immune system. Lactobacillus acidophilus, Lactobacillus reuteri, and Lactobacillus salivarius are members of the chicken intestinal microbiota and have been shown to induce different cytokine profiles in mononuclear cells in vitro. The objective of the present study was to examine the effects of these bacteria individually or in combination on the induction of antibody- and cell-mediated immune responses in vivo. The birds received lactobacilli weekly via oral gavage starting on day of hatch and subsequently, at 14 and 21 days, were immunized with sheep red blood cells (SRBC), keyhole limpet hemocyanin (KLH), Newcastle disease virus vaccine, and infectious bursal disease virus vaccine. Antibody responses in serum were measured weekly for 4 weeks beginning on the day of primary immunization. The cell-mediated immune response was evaluated at 21 days postimmunization by measurement of gamma interferon (IFN-γ) production in splenocytes stimulated with inactivated vaccine antigens. L. salivarius-treated birds had significantly more serum antibody to SRBC and KLH than birds that were not treated with probiotics. L. salivarius-treated birds also had decreased cell-mediated immune responses to recall antigen stimulation. L. reuteri treatment did not significantly affect the systemic immune response, while L. acidophilus treatment increased the antibody response to KLH. These results indicate that systemic antibody- and cell-mediated immune responses can be modulated by oral treatment with lactobacilli but that these bacteria may vary in their ability to modulate the immune response.

In vivo administration of ligands for chicken toll-like receptors 4 and 21 induces the expression of immune system genes in the spleen.

Toll-like receptors (TLRs) are a group of conserved proteins that play an important role in pathogen recognition in addition to the initiation and regulation of innate and adaptive immune responses. To date, several TLRs have been identified in chickens, each recognizing different ligands. TLR stimulation in chickens has been shown to play a role in host-responses to pathogens. However, the mechanisms through which TLRs modulate the chicken immune system have not been well examined. The present study was conducted to characterize the kinetics of responses to TLR4 and TLR21 stimulation in chickens following intramuscular injections of their corresponding ligands, lipopolysaccharide (LPS) and CpG oligodeoxynucleotides (ODNs), respectively. To this end, relative expression of cytokine genes in the spleen was determined at 2, 6, 12 and 24 h after injection of TLR ligands. The results indicated that LPS strongly induced the up-regulation of some immune system genes early on in the response to treatment, including interferon (IFN)-γ, interleukin (IL)-10, and IL-1ß. Furthermore, treatment with CpG ODN promoted the up-regulation of major histocompatibility complex (MHC)-II, IFN-γ and IL-10. The response to CpG ODN appeared to be somewhat delayed compared to the response to LPS. Moreover, we found a significant increase in IFN-α gene expression in response to LPS but not CpG ODNs. Future studies may be aimed to further characterize the molecular mechanisms of TLR activation in chickens or to exploit TLR agonists as vaccine adjuvants.

Marek's disease virus influences the expression of genes associated with IFN-gamma-inducible MHC class II expression.

Chickens infected with Marek's disease virus (MDV) become lifelong carriers regardless of their susceptibility to clinical disease. Therefore various viral immune-evasive mechanisms must play a role in MDV-host interactions. MDV has previously been shown to influence the expression of major histocompatibility complex (MHC) class II molecules. However, little is known about the underlying mechanisms of this phenomenon. In the present study, we studied the effect of MDV infection on the expression of several genes associated with IFN-gamma-inducible MHC class II expression at 4, 7, 14, and 21 days post-infection (dpi). There was a significant (p