Cathelicidin host defence peptides in cetartiodactyls \u2013 identification and characterization of novel members and modification for exploitation as anti-infective agents

Cathelicidins are a family of innate immunity effectors present in most vertebrates. Cathelicidin-derived antimicrobial peptides play an essential role in host defence, with a direct antimicrobial action as part of the innate immune responses and the capacity to modulate cellular immunity, affecting both innate and adaptive responses. Some can also have protective roles connected with wound healing, angiogenesis or sepsis. Cathelicidins are characterized by a conserved pro-region, known as 'cathelin-like' domain, carrying a highly variable, C-terminal antimicrobial domain. Many mammalian species express only one cathelicidin, but artiodactyles are known to express up to a dozen, bearing structurally and functionally very diverse AMPs. These including small disulfide-stabilized cyclic peptides, extended linear peptides rich in specific residues (e.g. proline, arginine, phenylalanine, tryptophan) and helical peptides of different sizes. Cetaceans are phylogenetic with artiodactyls, so are also expected to express this wide repertoire of cathelicidin-derived AMPs, whose functional characteristics may however have been shaped by the particular pathogenic microbiota they face in their aquatic lifestyle. With the known artiodactyl peptides, these might provide interesting leads for biomedical application. My Ph.D. followed to two parallel lines of research: i) the study of cathelicidin AMPs identified in the dolphin Tursiops truncatus (bottlenose dolphin), identified by searching genomic databases, to define their antimicrobial activities and obtain information on the modes-of-action, using different types of functional assays. These studies were carried out in parallel with selected orthologues from other artiodactyls, such as cow, sheep or pig, to screen for differences among them, and revealed both convergences and some interesting differences, especially relating to a proline-rich (PR-AMP) and an \uf061-helical one. These studies established a potent antimicrobial activity for the helical peptide, with a remarkable capacity too inhibit bacterial growth also at sub-lethal concentrations. The PR-AMP instead showed an increased internalization capacity, due to its apparent efficient use of alternative transporters. These data provided interesting insights into specific aspects of Cetaceans cathelicidins in relation to those of other mammals, and lay the groundwork for the possible development of novel antimicrobial agents. ii) the possible application of the bovine cathelicidin-derived PR-AMP Bac7(1-35) as a vehicle for delivering antibiotic cargo into susceptible bacterial cells. This peptide acts intracellularly, entering cells using specific transport systems. The aim was to conjugate it to the antibiotic tobramycin and expand its capacity to penetrate into Gram-negative pathogens, possibly overcoming resistance mechanisms. I began to develop synthetic strategies for modifying the antibiotic and then linking it to the peptide vehicle. Functional assays showed that one type of construct had a broader spectrum of activity than the individual components, and in some cases was active against resistant strains. This new strategy may potentially be applied to other types or classes of currently available antibiotics, as long as they can be modified for conjugation without affecting activity

Management of bacterial infection in the liver transplant candidate

Bacterial infection (BI) is a common cause of impairment of liver function in patients with cirrhosis, especially in the liver transplant candidates. These patients share an immunocompromised state and increased susceptibility to develop community and hospital-acquired infections. The changing epidemiology of BI, with an increase of multidrug resistant strains, especially in healthcare-associated settings, represents a critical issue both in the waiting list and in the post-operative management. This review focused on the role played by BI in patients awaiting liver transplantation, evaluating the risk of drop-out from the waiting list, the possibility to undergo liver transplantation after recovery from infection or during a controlled infection

Designing New Hybrid Antibiotics: Proline-Rich Antimicrobial Peptides Conjugated to the Aminoglycoside Tobramycin

Resistance to aminoglycoside antibiotics is a serious problem,typically arising from inactivating enzymes, reduced uptake, or increasedefflux in the important pathogens for which they are used as treatment.Conjugating aminoglycosides to proline-rich antimicrobial peptides(PrAMPs), which also target ribosomes and have a distinct bacterialuptake mechanism, might mutually benefit their individual activities.To this aim we have developed a strategy for noninvasively modifyingtobramycin to link it to a Cys residue and through this covalentlylink it to a Cys-modified PrAMP by formation of a disulfide bond.Reduction of this bridge in the bacterial cytosol should release theindividual antimicrobial moieties. We found that the conjugation oftobramycin to the well-characterized N-terminal PrAMP fragment Bac7(1-35)resulted in a potent antimicrobial capable of inactivating not onlytobramycin-resistant bacterial strains but also those less susceptibleto the PrAMP. To a certain extent, this activity also extends to theshorter and otherwise poorly active fragment Bac7(1-15). Althoughthe mechanism that allows the conjugate to act when its individualcomponents do not is as yet unclear, results are very promising andsuggest this may be a way of resensitizing pathogens that have developedresistance to the antibiotic

Myticalins: A novel multigenic family of linear, cationic antimicrobial peptides from marine mussels (Mytilus spp.)

The application of high-throughput sequencing technologies to non-model organisms has brought new opportunities for the identification of bioactive peptides from genomes and transcriptomes. From this point of view, marine invertebrates represent a potentially rich, yet largely unexplored resource for de novo discovery due to their adaptation to diverse challenging habitats. Bioinformatics analyses of available genomic and transcriptomic data allowed us to identify myticalins, a novel family of antimicrobial peptides (AMPs) from the mussel Mytilus galloprovincialis, and a similar family of AMPs from Modiolus spp., named modiocalins. Their coding sequence encompasses two conserved N-terminal (signal peptide) and C-terminal (propeptide) regions and a hypervariable central cationic region corresponding to the mature peptide. Myticalins are taxonomically restricted to Mytiloida and they can be classified into four subfamilies. These AMPs are subject to considerable interindividual sequence variability and possibly to presence/absence variation. Functional assays performed on selected members of this family indicate a remarkable tissue-specific expression (in gills) and broad spectrum of activity against both Gram-positive and Gram-negative bacteria. Overall, we present the first linear AMPs ever described in marine mussels and confirm the great potential of bioinformatics tools for the de novo discovery of bioactive peptides in non-model organisms

Direct-acting antivirals and hepatocellular carcinoma in chronic hepatitis C: A few lights and many shadows

With the introduction of direct-acting antiviral agents (DAA), the rate of sustained virological response (SVR) in the treatment of hepatitis C virus (HCV) has radically improved to over 95%. Robust scientific evidence supports a beneficial role of SVR after interferon therapy in the progression of cirrhosis, resulting in a decreased incidence of hepatocellular carcinoma (HCC). However, a debate on the impact of DAAs on the development of HCC is ongoing. This review aimed to analyse the scientific literature regarding the risk of HCC in terms of its recurrence and occurrence after the use of DAAs to eradicate HCV infection. Among 11 studies examining HCC occurrence, the de novo incidence rate ranged from 0 to 7.4% (maximum follow-up: 18 mo). Among 18 studies regarding HCC recurrence, the rate ranged from 0 to 54.4% (maximum "not well-defined" followup: 32 mo). This review highlights the major difficulties in interpreting data and reconciling the results of the included studies. These difficulties include heterogeneous cohorts, potential misclassifications of HCC prior to DAA therapy, the absence of an adequate control group, short follow-up times and different kinds of follow-up. Moreover, no clinical feature-based scoring system accounts for the molecular characteristics and pathobiology of the tumours. Nonetheless, this review does not suggest that there is a higher rate of de novo HCC occurrence or recurrence after DAA therapy in patients with previous HCV infection. \ua9 2018 The Author(s). Published by Baishideng Publishing Group Inc. All rights reserved

Assessing the impact of COVID-19 on liver cancer management (CERO-19).

BACKGROUND & AIMS: The coronavirus disease 2019 (COVID-19) pandemic has posed unprecedented challenges to healthcare systems and it may have heavily impacted patients with liver cancer (LC). Herein, we evaluated whether the schedule of LC screening or procedures has been interrupted or delayed because of the COVID-19 pandemic. METHODS: An international survey evaluated the impact of the COVID-19 pandemic on clinical practice and clinical trials from March 2020 to June 2020, as the first phase of a multicentre, international, and observational project. The focus was on patients with hepatocellular carcinoma or intrahepatic cholangiocarcinoma, cared for around the world during the first COVID-19 pandemic wave. RESULTS: Ninety-one centres expressed interest to participate and 76 were included in the analysis, from Europe, South America, North America, Asia, and Africa (73.7%, 17.1%, 5.3%, 2.6%, and 1.3% per continent, respectively). Eighty-seven percent of the centres modified their clinical practice: 40.8% the diagnostic procedures, 80.9% the screening programme, 50% cancelled curative and/or palliative treatments for LC, and 41.7% modified the liver transplantation programme. Forty-five out of 69 (65.2%) centres in which clinical trials were running modified their treatments in that setting, but 58.1% were able to recruit new patients. The phone call service was modified in 51.4% of centres which had this service before the COVID-19 pandemic (n = 19/37). CONCLUSIONS: The first wave of the COVID-19 pandemic had a tremendous impact on the routine care of patients with liver cancer. Modifications in screening, diagnostic, and treatment algorithms may have significantly impaired the outcome of patients. Ongoing data collection and future analyses will report the benefits and disadvantages of the strategies implemented, aiding future decision-making. LAY SUMMARY: The coronavirus disease 2019 (COVID-19) pandemic has posed unprecedented challenges to healthcare systems globally. Herein, we assessed the impact of the first wave pandemic on patients with liver cancer and found that routine care for these patients has been majorly disrupted, which could have a significant impact on outcomes

Energy consumption of high-performance buildings: Design vs. Reality

Studying the performance of highly efficient buildings is crucial for future policy, for example to inform decision making about whether to pursue further thermal improvement of buildings (insulation) or to rather foster investments in renewable resources. For this purpose, reliable values of the energy demand of new and retrofitted buildings are needed. However, there is evidence of a significant Energy Performance Gap (EPG) in buildings, defined as the difference between measured and calculated energy consumption. The objective of this paper is therefore to quantify the EPG in high efficiency buildings in Switzerland. The EPG was studied for 56 residential buildings, including various construction standards (Minergie, Minergie-P, and Minergie-A) and building types (new and retrofitted). The Minergie indexes were used as theoretical consumption, thereby representing the total final energy consumption for all needs of the buildings. These values were compared to data based on measurements. For the buildings in the sample the analysis yields a negative EPG of -14% (i.e. the median building consumes slightly less than its standard), indicating that the most efficient buildings are more robust to the EPG. However, this finding could be partly a consequence of the small sample used and its characteristics

GAPxPLORE: Energy Performance Gap in existing, new, and renovated buildings: Learning from large-scale datasets

The GAPxPLORE project aims to evaluate the EPG in the Swiss building stock. It studies how the EPG is distributed among the building typologies (performance level and age). Large-scale datasets from a range of sources containing calculated and real energy consumption data of buildings are analysed. GAPxPLORE also investigates the Energy Savings Deficit (ESD), defined as the difference between expected and achieved energy consumption reductions in renovated buildings. The focus lies on the final energy consumed for heating and domestic hot water in residential buildings

Induced expression of cathelicidins in trout (Oncorhynchus mykiss) challenged with four different bacterial pathogens

Cathelicidins are an important family of antimicrobial peptide effectors of innate immunity in vertebrates. Two members of this group, CATH\u20101 and CATH\u20102, have been identified and characterized in teleosts (ray\u2010finned fish). In this study, we investigated the expression of these genes in different tissues of rainbow trout challenged with 4 different inactivated pathogens. By using qPCR, we detected a strong induction of both cath\u20101 and cath\u20102 genes within 24 hours after intraperitoneal inoculation with Lactococcus garvieae, Yersinia ruckeri, Aeromonas salmonicida, or Flavobacterium psychrophilum cells. Up to 700\u2010fold induction of cath\u20102 was observed in the spleen of animals challenged with Y. ruckeri. Moreover, we found differences in the intensity and timing of gene up\u2010regulation in the analyzed tissues. The overall results highlight the importance of cathelicidins in the immune response mechanisms of salmonids