Publications by authors named "Chiara Pagliuca"

40 Publications

A novel smaller β-defensin-derived peptide is active against multidrug-resistant bacterial strains.

FASEB J 2021 12;35(12):e22026

Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, Naples, Italy.

Antibiotic resistance is becoming a severe obstacle in the fight against acute and chronic infectious diseases that accompany most degenerative illnesses from neoplasia to osteo-arthritis and obesity. Currently, the race is on to identify pharmaceutical molecules or combinations of molecules able to prevent or reduce the insurgence and/or progression of infectivity. Attempts to substitute antibiotics with antimicrobial peptides have, thus far, met with little success against multidrug-resistant (MDR) bacterial strains. During the last decade, we designed and studied the activity and features of human β-defensin analogs, which are salt-resistant, and hence active also under high salt concentrations as, for instance, in cystic fibrosis. Herein, we describe the design, synthesis, and major features of a new 21 aa long molecule, peptide γ2. The latter derives from the γ-core of the β-defensin natural molecules, a small fragment of these molecules still bearing high antibacterial activity. We found that peptide γ2, which contains only one disulphide bond, recapitulates most of the biological properties of natural human β-defensins and can also counteract both Gram-positive and Gram-negative MDR bacterial strains and biofilm formation. Moreover, it has great stability in human serum thereby enhancing its antibacterial presence and activity without cytotoxicity in human cells. In conclusion, peptide γ2 is a promising new weapon also in the battle against intractable infectious diseases.
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http://dx.doi.org/10.1096/fj.202002330RRDOI Listing
December 2021

Discovery and SAR Evolution of Pyrazole Azabicyclo[3.2.1]octane Sulfonamides as a Novel Class of Non-Covalent -Acylethanolamine-Hydrolyzing Acid Amidase (NAAA) Inhibitors for Oral Administration.

J Med Chem 2021 09 1;64(18):13327-13355. Epub 2021 Sep 1.

D3-PharmaChemistry, Istituto Italiano di Tecnologia (IIT), 16163Genova, Italy.

Inhibition of intracellular -acylethanolamine-hydrolyzing acid amidase (NAAA) activity is a promising approach to manage the inflammatory response under disabling conditions. In fact, NAAA inhibition preserves endogenous palmitoylethanolamide (PEA) from degradation, thus increasing and prolonging its anti-inflammatory and analgesic efficacy at the inflamed site. In the present work, we report the identification of a potent, systemically available, novel class of NAAA inhibitors, featuring a pyrazole azabicyclo[3.2.1]octane structural core. After an initial screening campaign, a careful structure-activity relationship study led to the discovery of -ethoxymethyl-pyrazinyloxy-8-azabicyclo[3.2.1]octane-pyrazole sulfonamide (), which was found to inhibit human NAAA in the low nanomolar range (IC = 0.042 μM) with a non-covalent mechanism of action. In light of its favorable biochemical, in vitro and in vivo drug-like profile, sulfonamide could be regarded as a promising pharmacological tool to be further investigated in the field of inflammatory conditions.
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http://dx.doi.org/10.1021/acs.jmedchem.1c00575DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8474119PMC
September 2021

Microbiological Evaluation and Sperm DNA Fragmentation in Semen Samples of Patients Undergoing Fertility Investigation.

Genes (Basel) 2021 04 27;12(5). Epub 2021 Apr 27.

Department of Molecular Medicine and Medical Biotechnologies, University of Naples Federico II, via S. Pansini 5, 80131 Napoli, Italy.

Fifteen percent of male infertility is associated with urogenital infections; several pathogens are able to alter the testicular and accessory glands' microenvironment, resulting in the impairment of biofunctional sperm parameters. The purpose of this study was to assess the influence of urogenital infections on the quality of 53 human semen samples through standard analysis, microbiological evaluation, and molecular characterization of sperm DNA damage. The results showed a significant correlation between infected status and semen volume, sperm concentration, and motility. Moreover, a high risk of fragmented sperm DNA was demonstrated in the altered semen samples. Urogenital infections are often asymptomatic and thus an in-depth evaluation of the seminal sample can allow for both the diagnosis and therapy of infections while providing more indicators for male infertility management.
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http://dx.doi.org/10.3390/genes12050654DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8145398PMC
April 2021

Phytocompounds vs. Dental Plaque Bacteria: Effects of Myrtle and Pomegranate Polyphenolic Extracts Against Single-Species and Multispecies Oral Biofilms.

Front Microbiol 2020 5;11:592265. Epub 2020 Nov 5.

Department of Science and Technology, University of Sannio, Benevento, Italy.

In the last decades, resistant microbial infection rate has dramatically increased, especially infections due to biofilm-producing strains that require increasingly complex treatments and are responsible for the increased mortality percentages compared with other infectious diseases. Considering that biofilms represent a key factor for a wide range of chronic infections with high drug tolerance, the treatment of biofilm-causing bacterial infections represents a great challenge for the future. Among new alternative strategies to conventional antimicrobial agents, the scientific interest has shifted to the study of biologically active compounds from plant-related extracts with known antimicrobial properties, in order to also evaluate their antibiofilm activity. In this regard, the aim of this study has been to assess the antibiofilm activity of polyphenolic extracts from myrtle leaf and pomegranate peel against oral pathogens of dental plaque, an excellent polymicrobial biofilm model. In particular, the antibiofilm properties of myrtle and pomegranate extracts, also in binary combination, were highlighted. In addition to inhibiting the biofilm formation, the tested polyphenolic extracts have been proven to destroy both preformed single-species and multispecies biofilms formed by , , , and oral isolates, suggesting that the new natural sources are rich in promising compounds able to counteract biofilm-related infections.
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http://dx.doi.org/10.3389/fmicb.2020.592265DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7674652PMC
November 2020

The Role of Thermal Water in Chronic Skin Diseases Management: A Review of the Literature.

J Clin Med 2020 Sep 22;9(9). Epub 2020 Sep 22.

Department of Clinical Medicine and Surgery, University "Federico II" of Naples, Via Pansini 5, 80131 Naples, Italy.

The benefits of thermal water in different diseases have been known since ancient times. Over the past decades, a re-assessment of the use of mineral water for the treatment of several pathologic conditions has taken place around the world. Today, water therapy is being practiced in many countries that have a variety of mineral springs considerably different in their hydrogeologic origin, temperature, and chemical composition. Thermal water and balneotherapy offer several advantages: this approach needs no chemicals or potentially harmful drugs; there are almost no side effects during and after treatment, and there is a low risk to the patient's general health and well-being. However, it is difficult to evaluate the efficacy of this therapeutic approach in clinical practice due to the complexity of molecular mechanisms underlying its efficacy. Here we review the current knowledge of the chemical, immunological, and microbiological basis for therapeutic effects of thermal water with a specific focus on chronic inflammatory skin diseases. We also describe recent evidence of the major dermatologic diseases that are frequently treated by balneotherapy with a remarkable rate of success. Moreover, we discuss the potential role of balneotherapy either alone or as a complement to conventional medical treatments.
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http://dx.doi.org/10.3390/jcm9093047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7563194PMC
September 2020

Synergy of Polyphenolic Extracts From Honey, Myrtle and Pomegranate Against Oral Pathogens, and .

Front Microbiol 2020 24;11:1465. Epub 2020 Jul 24.

Department of Science and Technology, University of Sannio, Benevento, Italy.

The increasing incidence rate of oral diseases, the wide spread of antimicrobial resistance, and the adverse effects of conventional antibiotics mean alternative prevention and treatment options are needed to counteract oral pathogens. In this regard, our study aims to evaluate the antibacterial activity of polyphenolic extracts prepared from acacia honey, myrtle leaves, and pomegranate peel against cariogenic bacteria, such as and . The chemical-physical parameters of acacia honey and the RP-HPLC polyphenolic profile of pomegranate peel extract have been previously described in our studies, while the characterization of myrtle extract, performed by HPLC analysis, is reported here. All the extracts were used singly and in binary combinations to highlight any synergistic effects. Moreover, the extracts were tested in association with amoxicillin to evaluate their ability to reduce the effective dose of this drug The values of minimal inhibitory concentrations and minimal bactericidal concentrations have been used to quantitatively measure the antibacterial activity of the single extracts, while the fractional inhibitory concentration index has been considered as predictor of anticariogenic synergistic effects. Finally, a time-kill curve method allowed for the evaluation of the bactericidal efficacy of the combined extracts. The microbiological tests suggest that acacia honey, myrtle, and pomegranate extracts are able to inhibit the cariogenic bacteria, also with synergistic effects. This study provides useful and encouraging results for the use of natural extract combinations alone or in association with antibiotics (adjuvant therapy) as a valid alternative for the prevention and treatment of oral infectious diseases.
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http://dx.doi.org/10.3389/fmicb.2020.01465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7396681PMC
July 2020

Urinary Biomarkers: Diagnostic Tools for Monitoring Athletes' Health Status.

Int J Environ Res Public Health 2020 08 20;17(17). Epub 2020 Aug 20.

Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Via S. Pansini 5, 80131 Naples, Italy.

Acute or intense exercise is sometimes related to infections of the urinary tract. It can also lead to incorrect hydration as well as incorrect glomerular filtration due to the presence of high-molecular-weight proteins that cause damage to the kidneys. In this context, our study lays the foundations for the use of a urine test in a team of twelve male basketball players as a means of monitoring numerous biochemical parameters, including pH, specific weight, color, appearance, presence of bacterial cells, presence of squamous cells, leukocytes, erythrocytes, proteins, glucose, ketones, bilirubin, hemoglobin, nitrite, and leukocyte esterase, to prevent and/or treat the onset of pathologies, prescribe personalized treatments for each athlete, and monitor the athletes' health status.
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http://dx.doi.org/10.3390/ijerph17176065DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7503896PMC
August 2020

[Validation of surgical masks during COVID19 emergency: activities at the University of Napoli Federico II].

G Ital Med Lav Ergon 2020 06;42(2):73-81

CeSMA Centro Servizi Metrologici e Tecnologici Avanzati, Università degli Studi di Napoli Federico II, Corso Nicolangelo Protopisani, 80146 Napoli, Italy.

Summary: During COVID-19 pandemic crisis, Italian Government has approved Law Decree no. 18 of 17 march 2020, in which art. 15 allows enterprises to produce, import and commercialize surgical masks notwithstanding the current rules of product certification. It is just required that the interested enterprises send to the Italian National Institute of Health a selfcertification in which they declare the technical characteristics of the masks and that masks are produced according to the safety requirements. In this context, a technical-scientific unit was established at the University of Napoli Federico II to provide interested enterprises with state-of-the-art consultancy, testing and measurement services, adhering to rigorous scientific protocols. Characterization tests were carried out on 163 surgical masks and/or materials for their construction and they have enabled the identification of pre-screening criteria to simplify the procedure for evaluating surgical masks using methods for assessing the filtration efficiency of particles and aerosols. Based on experimental results, it has been observed that a filtration efficiency for particles with sizes larger that 650 nm (PFE>650) exceeding 35% might guarantees a bacterial filtration efficiency (BFE) higher than 95% while BFE values higher than 98% are obtained when the PFE>650 is larger than 40%. PFE measurement is extremely simpler with respect to BFE, the latter being time-consuming and requiring specific equipment and methods for its realization. Many tested materials have shown the capability to assure high filtration efficiencies but Spundonded-Meltblown-Spunbonded (SMS), that are layers of non-woven fabric with different weights of Meltblown, can simultaneously guarantee high particle filtration efficiencies with pressure drop values (breathability) in the limits to classify the surgical masks as Type II/IIR. In fact, the fabric products analyzed so far have not been able to simultaneously guarantee adequate BFE and breathability values. On the contrary, Spunbonds of adequate weights can virtually verify both requirements and accredit themselves as possible materials for the production of surgical masks, at least of Type I. Further studies are needed to verify the possibility of producing low-cost, reusable surgical masks that could meet the criteria of circular economy.
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June 2020

Methicillin-Resistant : Risk for General Infection and Endocarditis Among Athletes.

Antibiotics (Basel) 2020 Jun 18;9(6). Epub 2020 Jun 18.

Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Via S.Pansini 5, 80131 Naples, Italy.

The first studies on (SA) infections in athletes were conducted in the 1980s, and examined athletes that perform in close physical contact, with particular attention to damaged or infected skin. Recent studies have used molecular epidemiology to shed light on the transmission of SA in professional athletes. These studies have shown that contact between athletes is prolonged and constant, and that these factors influence the appearance of infections caused by SA. These results support the need to use sanitary measures designed to prevent the appearance of SA infections. The factors triggering the establishment of SA within professional sports groups are the nasal colonization of SA, contact between athletes and sweating. Hence, there is a need to use the most modern molecular typing methods to evaluate the appearance of cutaneous SA disease. This review aims to summarize both the current SA infections known in athletes and the diagnostic methods employed for recognition, pointing to possible preventive strategies and the factors that can act as a springboard for the appearance of SA and subsequent transmission between athletes.
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http://dx.doi.org/10.3390/antibiotics9060332DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7345113PMC
June 2020

HNP-1 and HBD-1 as Biomarkers for the Immune Systems of Elite Basketball Athletes.

Antibiotics (Basel) 2020 Jun 7;9(6). Epub 2020 Jun 7.

Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Via S. Pansini 5, 80131 Naples, Italy.

Acute or strenuous exercise is sometimes related to upper respiratory tract infections in athletes. Practicing intense and regular exercise can lead to incorrect activation of the immune system, causing athletes to be excluded from training programs and competitions. Defensins are small antimicrobial peptides that are part of the innate immune system and dynamically involved in several biological activities. In this study, we highlight the role of human defensins in competitive basketball athletes. In particular, we consider the behavior of alpha- and beta-defensins together with white blood cells in a cohort of players. Moreover, we focus our attention on cortisol, a physiological indicator of stress, and testosterone, both of which are human hormones involved in muscle metabolism. The free-testosterone/cortisol ratio is considered to be an indicator of overtraining among athletes. This paper provides an up-to-date information of the role of human defensins as self-defense molecules during a continuous stressor such as long-term exercise, and it recognizes them as potential markers of infection.
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http://dx.doi.org/10.3390/antibiotics9060306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7345027PMC
June 2020

Human Defensins: A Novel Approach in the Fight against Skin Colonizing a.

Antibiotics (Basel) 2020 04 21;9(4). Epub 2020 Apr 21.

Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II, Via S. Pansini 5, 80131 Naples, Italy.

is a microorganism capable of causing numerous diseases of the human skin. The incidence of skin infections reflects the conflict between the host skin's immune defenses and the virulence elements. Antimicrobial peptides (AMPs) are small protein molecules involved in numerous biological activities, playing a very important role in the innate immunity. They constitute the defense of the host's skin, which prevents harmful microorganisms from entering the epithelial barrier, including However, uses ambiguous mechanisms against host defenses by promoting colonization and skin infections. Our review aims to provide a reference collection on host-pathogen interactions in skin disorders, including infections and its resistance to methicillin (MRSA). In addition to these, we discuss the involvement of defensins and other innate immunity mediators (i.e., toll receptors, interleukin-1, and interleukin-17), involved in the defense of the host against the skin disorders caused by , and then focus on the evasion mechanisms developed by the pathogenic microorganism under analysis. This review provides the "state of the art" on molecular mechanisms underlying skin infection and the pharmacological potential of AMPs as a new therapeutic strategy, in order to define alternative directions in the fight against cutaneous disease.
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http://dx.doi.org/10.3390/antibiotics9040198DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7235756PMC
April 2020

Campylobacter jejuni bacteremia in Italian pediatric patients with acute lymphoblastic leukemia: Report of two cases.

New Microbiol 2020 Apr 19;43(2):96-98. Epub 2020 Apr 19.

Department of Clinical Pathology, Virology Unit, "San Giovanni di Dio e Ruggi d'Aragona Hospital", Salerno, Italy.

Infections caused by Campylobacter jejuni are rarely associated with extraintestinal complications. C. jejuni bacteremia is difficult to detect in patients with hematological malignancies undergoing chemotherapy where the choice of appropriate antibiotic treatment is extremely important. We report two cases of C. jejuni bacteremia in Italian pediatric patients affected by acute lymphoblastic leukemia (ALL). Agreeing with the most recent epidemiological data, both clinical isolates showed a typical phenotypic antimicrobial resistance patterns with combined resistance to ciprofloxacin and tetracycline. To our knowledge, this is the first report of C. jejuni isolation from the blood of ALL pediatric patients in Italy, and it provides important epidemiological information on this rare infection.
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April 2020

Setup of Quantitative PCR for Oral spp. Evaluation in Celiac Disease Diagnosis.

Diagnostics (Basel) 2019 Dec 26;10(1). Epub 2019 Dec 26.

Ceinge Biotecnologie Avanzate S. C. a R. L., 80131 Naples, Italy.

Coeliac disease (CD) is a multifactorial autoimmune disorder and gut dysbiosis contributes to its pathogenesis. We previously profiled by 16S rRNA sequencing duodenal and oropharyngeal microbiomes in active CD (a-CD), gluten-free diet (GFD) patients, and controls (CO) and found significantly higher levels of spp., with pro-inflammatory activities, in a-CD patients than in the other two groups. In this study, we developed a fast and simple qPCR-based method to evaluate the abundance of the oral spp. and the diagnostic performances of the test in CD diagnosis. The spp. abundances detected by quantitative PCR (qPCR) were: CO = 0.14, GFD = 0.15, a-CD = 2.08, showing a similar trend to those previously measured by next generation sequencing (NGS). In particular, spp. values obtained by both methods were significantly higher ( < 0.001) in a-CD than in the other two groups GFD and CO-the latter almost overlapping. We calculated by ROC curve analysis the threshold of 1.12 ng/μL of spp. to discriminate between CO+GFD and a-CD patients with 100% and 96.7% of diagnostic sensitivity and specificity, respectively. In conclusion, our data, if confirmed in other cohorts, suggest the q-PCR evaluation of oral spp. could be a fast and simple method to assess CD-associated dysbiosis for diagnostic purposes.
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http://dx.doi.org/10.3390/diagnostics10010012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7168164PMC
December 2019

Inducing Meningococcal Meningitis Serogroup C in Mice via Intracisternal Delivery.

J Vis Exp 2019 11 5(153). Epub 2019 Nov 5.

Department of Molecular Medicine and Medical Biotechnology, University of Naples Federico II; CEINGE - Advanced Biotechnology;

Neisseria meningitidis (meningococcus) is a narrow-host-range microorganism, globally recognized as the leading cause of bacterial meningitis. Meningococcus is a transient colonizer of human nasopharynx of approximately 10% of healthy subject. In particular circumstances, it acquires an invasive ability to penetrate the mucosal barrier and invades the bloodstream causing septicaemia. In the latest case, fulminating sepsis could arise even without the consequent development of meningitis. Conversely, bacteria could poorly multiply in the bloodstream, cross the blood brain barrier, reach the central nervous system, leading to fulminant meningitis. The murine models of bacterial meningitis represent a useful tool to investigate the host-pathogen interactions and to analyze the pathogenetic mechanisms responsible for this lethal disease. Although, several experimental model systems have been evaluated over the last decades, none of these were able to reproduce the characteristic pathological events of meningococcal disease. In this experimental protocol, we describe a detailed procedure for the induction of meningococcal meningitis in a mouse model based on the intracisternal inoculation of bacteria. The peculiar signs of human meningitis were recorded in the murine host through the assessment of clinical parameters (e.g., temperature, body weight), evaluation of survival rate, microbiological analysis and histological examination of brain injury. When using intracisternal (i.cist.) inoculum, meningococci complete delivery directly into cisterna magna, leading to a very efficient meningococcal replication in the brain tissue. A 1,000-fold increase of viable count of bacteria is observed in about 18 h. Moreover, meningococci are also found in the spleen, and liver of infected mice, suggesting that the liver may represent a target organ for meningococcal replication.
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http://dx.doi.org/10.3791/60047DOI Listing
November 2019

Celiac disease-associated Neisseria flavescens decreases mitochondrial respiration in CaCo-2 epithelial cells: Impact of Lactobacillus paracasei CBA L74 on bacterial-induced cellular imbalance.

Cell Microbiol 2019 08 20;21(8):e13035. Epub 2019 May 20.

CEINGE-Biotecnologie Avanzate SCarl, Naples, Italy.

We previously identified a Neisseria flavescens strain in the duodenum of celiac disease (CD) patients that induced immune inflammation in ex vivo duodenal mucosal explants and in CaCo-2 cells. We also found that vesicular trafficking was delayed after the CD-immunogenic P31-43 gliadin peptide-entered CaCo-2 cells and that Lactobacillus paracasei CBA L74 (L. paracasei-CBA) supernatant reduced peptide entry. In this study, we evaluated if metabolism and trafficking was altered in CD-N. flavescens-infected CaCo-2 cells and if any alteration could be mitigated by pretreating cells with L. paracasei-CBA supernatant, despite the presence of P31-43. We measured CaCo-2 bioenergetics by an extracellular flux analyser, N. flavescens and P31-43 intracellular trafficking by immunofluorescence, cellular stress by TBARS assay, and ATP by bioluminescence. We found that CD-N. flavescens colocalised more than control N. flavescens with early endocytic vesicles and more escaped autophagy thereby surviving longer in infected cells. P31-43 increased colocalisation of N. flavescens with early vesicles. Mitochondrial respiration was lower (P < .05) in CD-N. flavescens-infected cells versus not-treated CaCo-2 cells, whereas pretreatment with L. paracasei-CBA reduced CD-N. flavescens viability and improved cell bioenergetics and trafficking. In conclusion, CD-N. flavescens induces metabolic imbalance in CaCo-2 cells, and the L. paracasei-CBA probiotic could be used to correct CD-associated dysbiosis.
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http://dx.doi.org/10.1111/cmi.13035DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6618323PMC
August 2019

Virulence Traits of a Serogroup C Meningococcus and Isogenic Mutant, Defective in Surface-Exposed Sialic Acid, in a Murine Model of Meningitis.

Infect Immun 2019 04 25;87(4). Epub 2019 Mar 25.

Department of Molecular Medicine and Medical Biotechnology, Federico II University, Naples, Italy

In serogroup C , the () gene codes for an UDP--acetylglucosamine 2-epimerase that catalyzes the conversion of UDP--acetyl-α-d-glucosamine into -acetyl-d-mannosamine and UDP in the first step in sialic acid biosynthesis. This enzyme is required for the biosynthesis of the (α2→9)-linked polysialic acid capsule and for lipooligosaccharide (LOS) sialylation. In this study, we have used a reference serogroup C meningococcal strain and an isogenic knockout mutant to investigate the pathogenetic role of surface-exposed sialic acids in a model of meningitis based on intracisternal inoculation of BALB/c mice. Results confirmed the key role of surface-exposed sialic acids in meningococcal pathogenesis. The 50% lethal dose (LD) of the wild-type strain 93/4286 was about four orders of magnitude lower than that of the mutant. Compared to the wild-type strain, the ability of this mutant to replicate in brain and spread systemically was severely impaired. Evaluation of brain damage evidenced a significant reduction in cerebral hemorrhages in mice infected with the mutant in comparison with the levels in those challenged with the wild-type strain. Histological analysis showed the typical features of bacterial meningitis, including inflammatory cells in the subarachnoid, perivascular, and ventricular spaces especially in animals infected with the wild type. Noticeably, 80% of mice infected with the wild-type strain presented with massive bacterial localization and accompanying inflammatory infiltrate in the , indicating high tropism of meningococci exposing sialic acids toward this brain structure and a specific involvement of the in the mouse model of meningococcal meningitis.
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http://dx.doi.org/10.1128/IAI.00688-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6434112PMC
April 2019

Enrichment of semen culture in the diagnosis of bacterial prostatitis.

J Microbiol Methods 2018 11 26;154:124-126. Epub 2018 Oct 26.

Department of Clinical Pathology, Virology Unit, "San Giovanni di Dio e Ruggid'Aragona Hospital", Salerno, Italy.

The objective was to investigate the diagnostic accuracy of our microbiological protocol to simplify the evaluation of bacterial prostatitis in the clinical practice. Our findings show the possibility to apply our alternative enrichment semen culture method to detect prostatic bacterial infection with higher sensitivity than the gold standard M&S technique.
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http://dx.doi.org/10.1016/j.mimet.2018.10.016DOI Listing
November 2018

Oropharyngeal microbiome evaluation highlights Neisseria abundance in active celiac patients.

Sci Rep 2018 07 23;8(1):11047. Epub 2018 Jul 23.

Ceinge Biotecnologie Avanzate scarl, Naples, Italy.

We previously profiled duodenal microbiome in active (a-), gluten-free diet (GFD) celiac disease (CD) patients and controls finding higher levels of the Proteobacterium Neisseria flavescens in a-CD patients than in the other two groups. Here, we investigate the oropharyngeal microbiome in CD patients and controls to evaluate whether this niche share microbial composition with the duodenum. We characterized by 16S rRNA gene sequencing the oropharyngeal microbiome in 14 a-CD, 22 GFD patients and 20 controls. Bacteroidetes, Proteobacteria and Firmicutes differed significantly between the three groups. In particular, Proteobacteria abounded in a-CD and Neisseria species mostly accounted for this abundance (p < 0.001), whereas Bacteroidetes were more present in control and GFD microbiomes. Culture-based oropharyngeal microbiota analysis confirmed the greater abundance of Proteobacteria and of Neisseria species in a-CD. Microbial functions prediction indicated a greater metabolic potential for degradation of aminoacids, lipids and ketone bodies in a-CD microbiome than in control and GFD microbiomes, in which polysaccharide metabolism predominated. Our results suggest a continuum of a-CD microbial composition from mouth to duodenum. We may speculate that microbiome characterization in the oropharynx, which is a less invasive sampling than the duodenum, could contribute to investigate the role of dysbiosis in CD pathogenesis.
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http://dx.doi.org/10.1038/s41598-018-29443-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6056421PMC
July 2018

Antibacterial Activity of Pomegranate Juice and Peel Extracts on Cariogenic Bacteria.

Biomed Res Int 2017 25;2017:2152749. Epub 2017 Oct 25.

Department of Science and Technology, Sannio University, Via Port'arsa, No. 11, 82100 Benevento, Italy.

Aim: To evaluate the antimicrobial activity of hydroalcoholic extracts of pomegranate ( L.) peel and juice, against the microorganisms considered the main etiologic agents of dental caries.

Methods: The values of the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) were determined against Clarke ATCC® 25175™ strain and clinical isolate.

Results: Peel extracts inhibit effectively the growth and survival of ATCC 25175 strain and clinical isolate with MIC and MBC values of 10 g/l and 15 g/l, respectively. Furthermore, the pomegranate juice extract showed high inhibitory activity against ATCC 25175 strain with a MIC value of 25 g/l and a MBC value of 40 g/l, whereas, against , it has displayed a moderate inhibitory activity, with MIC and MBC values of 20 g/l and 140 g/l, respectively.

Conclusions: microbiological tests demonstrate that the hydroalcoholic extracts of pomegranate juice and peel are able to contrast the main cariogenic bacteria involved in tooth decay. Although being preliminary data, our results suggest that pomegranate polyphenolic compounds could represent a good adjuvant for the prevention and treatment of dental caries.
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http://dx.doi.org/10.1155/2017/2152749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5676346PMC
July 2018

The complete 12 Mb genome and transcriptome of Nonomuraea gerenzanensis with new insights into its duplicated "magic" RNA polymerase.

Sci Rep 2016 Dec 21;6(1):18. Epub 2016 Dec 21.

Department of Biological and Environmental Sciences and Technologies (DiSTeBA), University of Salento, Lecce, Italy.

In contrast to the widely accepted consensus of the existence of a single RNA polymerase in bacteria, several actinomycetes have been recently shown to possess two forms of RNA polymerases due the to co-existence of two rpoB paralogs in their genome. However, the biological significance of the rpoB duplication is obscure. In this study we have determined the genome sequence of the lipoglycopeptide antibiotic A40926 producer Nonomuraea gerenzanensis ATCC 39727, an actinomycete with a large genome and two rpoB genes, i.e. rpoB(S) (the wild-type gene) and rpoB(R) (the mutant-type gene). We next analyzed the transcriptional and metabolite profiles in the wild-type gene and in two derivative strains over-expressing either rpoB(R) or a mutated form of this gene to explore the physiological role and biotechnological potential of the "mutant-type" RNA polymerase. We show that rpoB(R) controls antibiotic production and a wide range of metabolic adaptive behaviors in response to environmental pH. This may give interesting perspectives also with regard to biotechnological applications.
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http://dx.doi.org/10.1038/s41598-016-0025-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5431353PMC
December 2016

Genotyping of Toxoplasma gondii strain directly from human CSF samples of congenital toxoplasmosis clinical case.

New Microbiol 2017 Apr 3;40(2):151-154. Epub 2017 Apr 3.

Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, Naples, Italy.

This report describes a case of congenital toxoplasmosis in a newborn in Southern Italy. A pregnant mother had been admitted at the 20th week of her pregnancy on account of pharyngodynia and laterocervical lymphadenopathy. Although serological testing of the mother's serum documented a seroconversion with positive IgG and IgM anti-Toxoplasma antibodies during II trimester, the woman refused to perform prenatal diagnosis for congenital toxoplasmosis. Fetal ultrasound scan already showed mild asymmetrical triventricular hydrocephaly and cerebral calcifications. After birth, real-time PCR on cerebrospinal fluid and blood samples of the newborn showed a positive result for 529bp-repeat element DNA of T. gondii, In addition brain magnetic resonance imaging and computed tomography showed a characteristic diffuse brain tissue loss associated with hydrocephalus. For the first time molecular characterization of T. gondii isolate was performed directly from the newborn's CSF samples by using nested-PCR-RFLP of sag-2 and pk1 genes. The PCR-RLFP analysis revealed that the isolate belongs to the clonal type II, the predominant lineage causing human toxoplasmosis, as confirmed by DNA sequencing.
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April 2017

Novel Approach for Evaluation of Protective Role against Liver Damage in Immunocompromised Murine Model.

Front Microbiol 2016 7;7:1750. Epub 2016 Nov 7.

Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical SchoolNaples, Italy; CEINGE-Advanced BiotechnologiesNaples, Italy.

is a gram-negative facultative intracellular bacterium and is the causative agent of cat-scratch disease. Our previous data have established that colonization is able to prevent damages through the polysaccharide A (PSA) in an experimental murine model. In order to determine whether the PSA is essential for the protection against pathogenic effects of in immunocompromised hosts, SCID mice were co-infected with wild type or its mutant ΔPSA and the effects of infection on murine tissues have been observed by High-Frequency Ultrasound (HFUS), histopathological examination, and Transmission Electron Microscopy (TEM). For the first time, echostructure, hepatic lobes length, vascular alterations, and indirect signs of hepatic dysfunctions, routinely used as signs of disease in humans, have been analyzed in an immunocompromised murine model. Our findings showed echostructural alterations in all infected mice compared with the Phosphate Buffer Solution (PBS) control group; further, those infected with and co-infected with ΔPSA presented the major echostructural alterations. Half of the mice infected with and all those co-infected with ΔPSA have showed an altered hepatic echogenicity compared with the renal cortex. The echogenicity score of co-infected mice with ΔPSA differed significantly compared with the PBS control group (p < 0.05). Moreover the inflammation score of the histopathological evaluation was fairly concordant with ultrasound findings. Ultrastructural analysis performed by TEM revealed no significant alterations in liver samples of SCID mice infected with wild type while those infected with ΔPSA showed the presence of collagen around the main vessels compared with the PBS control group. The liver samples of mice infected with showed macro-areas rich in collagen, stellate cells, and histiocytic cells. Interestingly, our data demonstrated that immunocompromised SCID mice infected with and co-infected with ΔPSA showed the most severe morpho-structural liver damage. In addition, these results suggests that the HFUS together with histopathological evaluation could be considered good imaging approach to evaluate hepatic alterations.
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http://dx.doi.org/10.3389/fmicb.2016.01750DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5097911PMC
November 2016

Metagenomics Reveals Dysbiosis and a Potentially Pathogenic N. flavescens Strain in Duodenum of Adult Celiac Patients.

Am J Gastroenterol 2016 Jun 5;111(6):879-90. Epub 2016 Apr 5.

CEINGE-Biotecnologie Avanzate, Naples, Italy.

Objectives: Celiac disease (CD)-associated duodenal dysbiosis has not yet been clearly defined, and the mechanisms by which CD-associated dysbiosis could concur to CD development or exacerbation are unknown. In this study, we analyzed the duodenal microbiome of CD patients.

Methods: The microbiome was evaluated in duodenal biopsy samples of 20 adult patients with active CD, 6 CD patients on a gluten-free diet, and 15 controls by DNA sequencing of 16S ribosomal RNA libraries. Bacterial species were cultured, isolated and identified by mass spectrometry. Isolated bacterial species were used to infect CaCo-2 cells, and to stimulate normal duodenal explants and cultured human and murine dendritic cells (DCs). Inflammatory markers and cytokines were evaluated by immunofluorescence and ELISA, respectively.

Results: Proteobacteria was the most abundant and Firmicutes and Actinobacteria the least abundant phyla in the microbiome profiles of active CD patients. Members of the Neisseria genus (Betaproteobacteria class) were significantly more abundant in active CD patients than in the other two groups (P=0.03). Neisseria flavescens (CD-Nf) was the most abundant Neisseria species in active CD duodenum. Whole-genome sequencing of CD-Nf and control-Nf showed genetic diversity of the iron acquisition systems and of some hemoglobin-related genes. CD-Nf was able to escape the lysosomal compartment in CaCo-2 cells and to induce an inflammatory response in DCs and in ex-vivo mucosal explants.

Conclusions: Marked dysbiosis and an abundance of a peculiar CD-Nf strain characterize the duodenal microbiome in active CD patients thus suggesting that the CD-associated microbiota could contribute to the many inflammatory signals in this disorder.
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http://dx.doi.org/10.1038/ajg.2016.95DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4897008PMC
June 2016

Potent α-amino-β-lactam carbamic acid ester as NAAA inhibitors. Synthesis and structure-activity relationship (SAR) studies.

Eur J Med Chem 2016 Mar 27;111:138-59. Epub 2016 Jan 27.

Drug Discovery and Development, Italian Institute of Technology, Via Morego 30, I-16163 Genova, Italy; Departments of Anatomy and Neurobiology, Pharmacology and Biological Chemistry, University of California, Irvine 92697-4625, USA. Electronic address:

4-Cyclohexylbutyl-N-[(S)-2-oxoazetidin-3-yl]carbamate (3b) is a potent, selective and systemically active inhibitor of intracellular NAAA activity, which produces profound anti-inflammatory effects in animal models. In the present work, we describe structure-activity relationship (SAR) studies on 3-aminoazetidin-2-one derivatives, which have led to the identification of 3b, and expand these studies to elucidate the principal structural and stereochemical features needed to achieve effective NAAA inhibition. Investigations on the influence of the substitution at the β-position of the 2-oxo-3-azetidinyl ring as well as on the effect of size and shape of the carbamic acid ester side chain led to the discovery of 3ak, a novel inhibitor of human NAAA that shows an improved physicochemical and drug-like profile relative to 3b. This favourable profile, along with the structural diversity of the carbamic acid chain of 3b, identify this compound as a promising new tool to investigate the potential of NAAA inhibitors as therapeutic agents for the treatment of pain and inflammation.
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http://dx.doi.org/10.1016/j.ejmech.2016.01.046DOI Listing
March 2016

Fitness Cost of Rifampin Resistance in Neisseria meningitidis: In Vitro Study of Mechanisms Associated with rpoB H553Y Mutation.

Antimicrob Agents Chemother 2015 Dec 28;59(12):7637-49. Epub 2015 Sep 28.

Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, Naples, Italy Ceinge Advanced Biotechnologies, Naples, Italy

Rifampin chemoprophylaxis against Neisseria meningitidis infections led to the onset of rifampin resistance in clinical isolates harboring point mutations in the rpoB gene, coding for the RNA polymerase β chain. These resistant strains are rare in medical practice, suggesting their decreased fitness in the human host. In this study, we isolated rifampin-resistant rpoB mutants from hypervirulent serogroup C strain 93/4286 and analyzed their different properties, including the ability to grow/survive in different culture media and in differentiated THP-1 human monocytes and to compete with the wild-type strain in vitro. Our results demonstrate that different rpoB mutations (H553Y, H553R, and S549F) may have different effects, ranging from low- to high-cost effects, on bacterial fitness in vitro. Moreover, we found that the S549F mutation confers temperature sensitivity, possibly explaining why it is observed very rarely in clinical isolates. Comparative high-throughput RNA sequencing analysis of bacteria grown in chemically defined medium demonstrated that the low-cost H553Y substitution resulted in global transcriptional changes that functionally mimic the stringent response. Interestingly, many virulence-associated genes, including those coding for meningococcal type IV pili, porin A, adhesins/invasins, IgA protease, two-partner secretion system HrpA/HrpB, enzymes involved in resistance to oxidative injury, lipooligosaccharide sialylation, and capsular polysaccharide biosynthesis, were downregulated in the H553Y mutant compared to their level of expression in the wild-type strain. These data might account for the reduced capacity of this mutant to grow/survive in differentiated THP-1 cells and explain the rarity of H553Y mutants among clinical isolates.
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http://dx.doi.org/10.1128/AAC.01746-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4649176PMC
December 2015

Seroprevalence of Bartonella henselae in patients awaiting heart transplant in Southern Italy.

J Microbiol Immunol Infect 2017 Apr 14;50(2):239-244. Epub 2015 May 14.

U.O.C. Division of Immunohematology, Transfusion Medicine and Transplant Immunology, Regional Reference Laboratory of Transplant Immunology, Azienda Ospedaliera Universitaria, Second University of Naples, Naples, Italy; Institute of Diagnostic and Nuclear Development (SDN), Istituto Di Ricovero e Cura a Carattere Scientifico (IRCCS), Naples, Italy.

Background: Bartonella henselae is the etiologic agent of cat-scratch disease. B. henselae infections are responsible for a widening spectrum of human diseases, although often symptomless, ranging from self-limited to life-threatening and show different courses and organ involvement due to the balance between host and pathogen. The role of the host immune response to B. henselae is critical in preventing progression to systemic disease. Indeed in immunocompromised patients, such as solid organ transplant patients, B. henselae results in severe disseminated disease and pathologic vasoproliferation. The purpose of this study was to determine the seroprevalence of B. henselae in patients awaiting heart transplant compared to healthy individuals enrolled in the Regional Reference Laboratory of Transplant Immunology of Second University of Naples.

Methods: Serum samples of 38 patients awaiting heart transplant in comparison to 50 healthy donors were examined using immunfluorescence assay.

Results: We found a B. henselae significant antibody positivity rate of 21% in patients awaiting heart transplant (p = 0.002). There was a positive rate of 8% (p > 0.05) for immunoglobulin (Ig)M and a significant value of 13% (p = 0.02) for IgG, whereas controls were negative both for IgM and IgG antibodies against B. henselae. The differences in comorbidity between cases and controls were statistically different (1.41 ± 0.96 vs 0.42 ± 0.32; p = 0.001).

Conclusions: Although this study was conducted in a small number of patients, we suggest that the identification of these bacteria should be included as a routine screening analysis in pretransplant patients.
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http://dx.doi.org/10.1016/j.jmii.2015.05.001DOI Listing
April 2017

Benzoxazolone carboxamides: potent and systemically active inhibitors of intracellular acid ceramidase.

Angew Chem Int Ed Engl 2015 Jan 13;54(2):485-9. Epub 2014 Nov 13.

Drug Discovery and Development, Istituto Italiano di Tecnologia, Via Morego 30, 16163 Genova (Italy).

The ceramides are a family of bioactive lipid-derived messengers involved in the control of cellular senescence, inflammation, and apoptosis. Ceramide hydrolysis by acid ceramidase (AC) stops the biological activity of these substances and influences survival and function of normal and neoplastic cells. Because of its central role in the ceramide metabolism, AC may offer a novel molecular target in disorders with dysfunctional ceramide-mediated signaling. Here, a class of benzoxazolone carboxamides is identified as the first potent and systemically active inhibitors of AC. Prototype members of this class inhibit AC with low nanomolar potency by covalent binding to the catalytic cysteine. Their metabolic stability and high in vivo efficacy suggest that these compounds may be used as probes to investigate the roles of ceramide in health and disease, and that this scaffold may represent a promising starting point for the development of novel therapeutic agents.
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http://dx.doi.org/10.1002/anie.201409042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4502975PMC
January 2015

Identification of Inquilinus limosus in cystic fibrosis: a first report in Italy.

New Microbiol 2014 Oct 1;37(4):567-71. Epub 2014 Oct 1.

Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, Naples, Italy.

Cystic fibrosis is a genetic disorder associated with a polymicrobial lung infection where classical pathogens and newly identified bacteria may interact. Inquilinus limosus is an a-proteobacterium recently isolated in the airways of cystic fibrosis patient. We report the first case in Italy of I.limosus isolation from the sputum sample of a cystic fibrosis patient. The patient is a 20-years-old man with cystic fibrosis, regularly attending the Regional Care Center for Cystic Fibrosis at the Federico II University Hospital of Naples. Microbiological culture methods detected a mu- coid gram negative bacillus in the patient's sputum sample. The isolate exhibited a distinct antimicrobial suscep- tibility profile with a high MIC for several drugs. The MALDI-TOF mass spectrometry analysis indicated the bac- terium isolated as I. limosus, confirmed by 16s rDNA sequence analysis. The described clinical case demonstrates how the bacterial biodiversity in the airways of cystic fibrosis patients is still underestimated. Cystic fibrosis lung represents an ecological niche suitable for growth of a wide variety of unusual bacteria not commonly associated with human diseases, such as I. limosus. Therefore further studies are needed to evaluate the epidemiology and clinical implications of I. limosus in the physiopathology of cystic fibrosis lung infection.
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October 2014
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