Nitrosative stress defences of the enterohepatic pathogenic bacterium Helicobacter pullorum

Helicobacter pullorum is an avian bacterium that causes gastroenteritis, intestinal bowel and hepatobiliary diseases in humans. Although H. pullorum has been shown to activate the mammalian innate immunity with release of nitric oxide (NO), the proteins that afford protection against NO and reactive nitrogen species (RNS) remain unknown. Here several protein candidates of H. pullorum, namely a truncated (TrHb) and a single domain haemoglobin (SdHb), and three peroxiredoxin-like proteins (Prx1, Prx2 and Prx3) were investigated. We report that the two haemoglobin genes are induced by RNS, and that SdHb confers resistance to nitrosative stress both in vitro and in macrophages. For peroxiredoxins, the prx2 and prx3 expression is enhanced by peroxynitrite and hydrogen peroxide, respectively. Mutation of prx1 does not alter the resistance to these stresses, while the single ∆prx2 and double ∆prx1∆prx2 mutants have decreased viability. To corroborate the physiological data, the biochemical analysis of the five recombinant enzymes was done, namely by stopped-flow spectrophotometry. It is shown that H. pullorum SdHb reacts with NO much more quickly than TrHb, and that the three Prxs react promptly with peroxynitrite, Prx3 displaying the highest reactivity. Altogether, the results unveil SdHb and Prx3 as major protective systems of H. pullorum against nitrosative stress

Cardiolipin, Mitochondria, and Neurological Disease

Over the past decade, it has become clear that lipid homeostasis is central to cellular metabolism. Lipids are particularly abundant in the central nervous system (CNS) where they modulate membrane fluidity, electric signal transduction, and synaptic stabilization. Abnormal lipid profiles reported in Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), and traumatic brain injury (TBI), are further support for the importance of lipid metablism in the nervous system. Cardiolipin (CL), a mitochondria-exclusive phospholipid, has recently emerged as a focus of neurodegenerative disease research. Aberrant CL content, structure, and localization are linked to impaired neurogenesis and neuronal dysfunction, contributing to aging and the pathogenesis of several neurodegenerative diseases, such as AD and PD. Furthermore, the highly tissue-specific acyl chain composition of CL confers it significant potential as a biomarker to diagnose and monitor the progression in several neurological diseases. CL also represents a potential target for pharmacological strategies aimed at treating neurodegeneration. Given the equipoise that currently exists between CL metabolism, mitochondrial function, and neurological disease, we review the role of CL in nervous system physiology and monogenic and neurodegenerative disease pathophysiology, in addition to its potential application as a biomarker and pharmacological target

The Lepidopteran endoribonuclease-U domain protein P102 displays dramatically reduced enzymatic activity and forms functional amyloids

Hemocytes of Heliothis virescens (F.) (Lepidoptera, Noctuidae) larvae produce a protein, P102, with a putative endoribonuclease-U domain. In previous works we have shown that P102 is involved in Lepidopteran immune response by forming amyloid fibrils, which catalyze and localize melanin deposition around non-self intruders during encapsulation, preventing harmful systemic spreading. Here we demonstrate that P102 belongs to a new class of proteins that, at least in Lepidoptera, has a diminished endoribonuclease-U activity probably due to the lack of two out of five catalytically essential residues. We show that the P102 homolog from Trichoplusia ni (Lepidoptera, Noctuidae) displays catalytic site residues identical to P102, a residual endoribonuclease-U activity and the ability to form functional amyloids. On the basis of these results as well as sequence and structural analyses, we hypothesize that all the Lepidoptera endoribonuclease-U orthologs with catalytic site residues identical to P102 form a subfamily with similar function

A mobile black soldier fly farm for on-site disposal of animal dairy manure

Black soldier fly (BSF), Hermetia illucens (L.) (Diptera Stratiomyidae), is a saprophagous insect that is receiving a growing scientific and economic interest since during the larval stage it is extremely voracious and able to consume a wide range of organic materials. This ethological characteristic is particularly suitable for waste management at industrial scale. The extraordinary ability to accumulate high levels of proteins and lipids, allows the use of resulting larvae as animal feed or biodiesel production; the residue of the bioconversion process, that consists of larval frass and not converted organic matter is assimilable to organic fertilizer. The aim of this study was to evaluate the bioconversion process by black soldier fly larvae (BSFL) on fresh and mature dairy manure. A “mobile bioconversion unit” that works as a mobile breeding unit was used, allowing to carry out bioconversion tests directly on fields, in different livestock farms located on the Basilicata territory (Italy). Total larval and frass biomass, bioconversion yield, BSFL development time and substrate reduction were evaluated for each treatment. All the analysed parameters differed from the control (larvae fed in standard diet) but not between the two substrates from the zootechnical chain. Although development time significantly differed across treatments, BSF correctly grow and reduce all substrates confirming larvae can be used to bioconvert animal manure reducing the undesired effects occurring from mismanaged this kind of substrate

Amyloid/Melanin distinctive mark in invertebrate immunity

Protostomes and Deuterostomes show the same nexus between melanin production, and amyloid fibril production, i.e., the presence of melanin is indissolubly linked to amyloid scaffold that, in turn, is conditioned by the redox status/cytoplasmic pH modification, pro-protein cleavage presence, adrenocorticotropin hormone (ACTH), melanocyte-stimulating hormone (\u3b1-MSH), and neutral endopeptidase (NEP) overexpressions. These events represent the crucial component of immune response in invertebrates, while in vertebrates these series of occurrences could be interpreted as a modest and very restricted innate immune response. On the whole, it emerges that the mechanisms involving amyloid fibrils/pigment synthesis in phylogenetically distant metazoan (viz, cnidaria, molluscs, annelids, insects, ascidians and vertebrates) are evolutionary conserved. Furthermore, our data show the relationship between immune and neuroendocrine systems in amyloid/melanin synthesis. Indeed the process is closely associated to ACTH-\u3b1-MSH production, and their role in stress responses leading to pigment production reflects and confirms again their ancient phylogeny

G-Quadruplex Dynamics Contribute To Regulation Of Mitochondrial Gene Expression

Single-stranded DNA or RNA sequences rich in guanine (G) can adopt non-canonical structures known as G-quadruplexes (G4). Mitochondrial DNA (mtDNA) sequences that are predicted to form G4 are enriched on the heavy-strand and have been associated with formation of deletion breakpoints. Increasing evidence supports the ability of mtDNA to form G4 in cancer cells; however, the functional roles of G4 structures in regulating mitochondrial nucleic acid homeostasis in non-cancerous cells remain unclear. Here, we demonstrate by live cell imaging that the G4-ligand RHPS4 localizes primarily to mitochondria at low doses. We find that low doses of RHPS4 do not induce a nuclear DNA damage response but do cause an acute inhibition of mitochondrial transcript elongation, leading to respiratory complex depletion. We also observe that RHPS4 interferes with mtDNA levels or synthesis both in cells and isolated mitochondria. Importantly, a mtDNA variant that increases G4 stability and anti-parallel G4-forming character shows a stronger respiratory defect in response to RHPS4, supporting the conclusion that mitochondrial sensitivity to RHPS4 is G4-mediated. Taken together, our results indicate a direct role for G4 perturbation in mitochondrial genome replication, transcription processivity, and respiratory function in normal cells

Insect antimicrobial peptides: potential weapons to counteract the antibiotic resistance

Misuse and overuse of antibiotics have contributed in the last decades to a phenomenon known as antibiotic resistance which is currently considered one of the principal threats to global public health by the World Health Organization. The aim to find alternative drugs has been demonstrated as a real challenge. Thanks to their biodiversity, insects represent the largest class of organisms in the animal kingdom. The humoral immune response includes the production of antimicrobial peptides (AMPs) that are released into the insect hemolymph after microbial infection. In this review, we have focused on insect immune responses, particularly on AMP characteristics, their mechanism of action and applications, especially in the biomedical field. Furthermore, we discuss the Toll, Imd, and JAK-STAT pathways that activate genes encoding for the expression of AMPs. Moreover, we focused on strategies to improve insect peptides stability against proteolytic susceptibility such as D-amino acid substitutions, N-terminus modification, cyclization and dimerization

Lipids from hermetia illucens, an innovative and sustainable source

The exponential increase of global demand for proteins and lipids can no longer be satisfied by classical sources. High amounts of CO2 produced by intensive livestock breeding and its effects on the environment are the main factors that prevent the use of animals as primary sources for proteins and lipids, calling for the use of new sustainable sources, such as insects. The massive breeding of bioconverter insects as a feed source has been a major topic in recent years, with both economic and scientific aspects related to rearing and subsequent processing optimization. The larvae of Hermetia illucens (Diptera: Stratiomyidae) (also known as Black Soldier Fly) can be used for the eco-sustainable production of proteins and lipids with high biological and economic value. Lipids can be obtained from BSF bioconversion processes and are present in high quantities in the last instar larvae and prepupae. Fats obtained from BSF are used as animal feed ingredients, in the formulation of several products for personal care, and in biodiesel production. To enable the use of insect-derived lipids, it is important to understand how to optimize their extraction. Here, we summarize the published information on the composition, the extraction methods, and the possible applications of the BSF lipid component

In Vitro Evaluation of the Antibacterial Activity of the Peptide Fractions Extracted from the Hemolymph of Hermetia illucens (Diptera: Stratiomyidae)

Antimicrobial peptides (AMPs) are a chemically and structurally heterogeneous family of molecules produced by a large variety of living organisms, whose expression is predominant in the sites most exposed to microbial invasion. One of the richest natural sources of AMPs is insects which, over the course of their very long evolutionary history, have adapted to numerous and different habitats by developing a powerful innate immune system that has allowed them to survive but also to assert themselves in the new environment. Recently, due to the increase in antibiotic-resistant bacterial strains, interest in AMPs has risen. In this work, we detected AMPs in the hemolymph of Hermetia illucens (Diptera, Stratiomyidae) larvae, following infection with Escherichia coli (Gram negative) or Micrococcus flavus (Gram positive) and from uninfected larvae. Peptide component, isolated via organic solvent precipitation, was analyzed by microbiological techniques. Subsequent mass spectrometry analysis allowed us to specifically identify peptides expressed in basal condition and peptides differentially expressed after bacterial challenge. We identified 33 AMPs in all the analyzed samples, of which 13 are specifically stimulated by Gram negative and/or Gram positive bacterial challenge. AMPs mostly expressed after bacterial challenge could be responsible for a more specific activity