Giorgia Borriello

Scopus Publications

The Use of Antimicrobials in Animal Husbandry as a Potential Factor for the Increased Incidence of Colorectal Cancer: Food Safety and Kinetics in a Murine Model
Rosa D’Ambrosio, Stefania Cavallo, Roberta Brunetti, Roberta Pellicanò, Emanuela Vaccaro, Giorgia Borriello, Rubina Paradiso, Francesco Paolo Serpe, Sara Lambiase, Francesca Bruzzese,et al.
MDPI AG
The aim of this research was to investigate the effects of the prolonged use of the broad-spectrum antimicrobial widely used in animal husbandry. By means of a mouse model, a translational study was carried out on immunocompetent mice (with a complete immune system). This study highlighted the effect of antimicrobial residues taken in with food on the growth time of cancer and on alterations to the gut microbiota. This project considered the fight against antimicrobial resistance from a One Health perspectivethrough collaboration between human medicine and veterinary medicine. Regarding food safety, antimicrobial residues in products of animal origin are rarely detected; they therefore constitute a negligible factor in determining colorectal cancer.

Streptococcus suis-associated neonatal meningitis and sepsis: characterization, antimicrobial resistance, and public health implications
Giovanna Fusco, Rubina Paradiso, Lorena Cardillo, Maria Antonia Salvia, Saveria Dodaro, Veronica Del Monaco, Gianfranco Scarpelli, Francesca Greco, Antonio Rinaldi, Lorella Barca,et al.
Frontiers Media SA
Neonatal meningitis and sepsis were diagnosed in a 32-day-old preterm-born infant. Streptococcus (S.) suis was isolated from cerebrospinal fluid and blood. Next-generation sequencing revealed that the strain was serotype 2 sequence type 1, and contained the ermB and tet(W) genes, which are responsible for resistance to macrolides and tetracycline, along with several pilus-associated genes and 20 virulence factors. High homology was observed with previously identified human and swine strains in the same area. S. suis meningitis and sepsis are mainly reported in adults, related to direct contact with pigs or contaminated pork meat consumption, while it is rarely reported in children. Herein, we describe the first case of S. suis in a newborn associated with meningitis and antimicrobial resistance. The rates of resistance to tetracyclines, lincosamides, and macrolides for this bacterium are increasing and are creating concern worldwide. Altogether, our findings highlight the importance of investigating S. suis in cases of neonatal meningitis, as well as the necessity of assessing the antimicrobial profile to obtain useful information for developing targeted therapies.

Genetic diversity of Brucella abortus strains from cattle and water buffalo in the Italian province of Caserta
Luigi Orrù, Antonella Lamontanara, Celestina Mascolo, Giorgia Borriello, Rubina Paradiso, Anna Cerrone, Paolo Coppa, Manuela Tittarelli, Carlo Ferrara, Esterina De Carlo,et al.
Elsevier BV

Exposure to environmental pollutants selects for xenobiotic-degrading functions in the human gut microbiome
Francesca De Filippis, Vincenzo Valentino, Giuseppina Sequino, Giorgia Borriello, Marita Georgia Riccardi, Biancamaria Pierri, Pellegrino Cerino, Antonio Pizzolante, Edoardo Pasolli, Mauro Esposito,et al.
Springer Science and Business Media LLC
AbstractEnvironmental pollutants from different chemical families may reach the gut microbiome, where they can be metabolized and transformed. However, how our gut symbionts respond to the exposure to environmental pollution is still underexplored. In this observational, cohort study, we aim to investigate the influence of environmental pollution on the gut microbiome composition and potential activity by shotgun metagenomics. We select as a case study a population living in a highly polluted area in Campania region (Southern Italy), proposed as an ideal field for exposomic studies and we compare the fecal microbiome of 359 subjects living in areas with high, medium and low environmental pollution. We highlight changes in gut microbiome composition and functionality that were driven by pollution exposure. Subjects from highly polluted areas show higher blood concentrations of dioxin and heavy metals, as well as an increase in microbial genes related to degradation and/or resistance to these molecules. Here we demonstrate the dramatic effect that environmental xenobiotics have on gut microbial communities, shaping their composition and boosting the selection of strains with degrading capacity. The gut microbiome can be considered as a pivotal player in the environment-health interaction that may contribute to detoxifying toxic compounds and should be taken into account when developing risk assessment models. The study was registered at ClinicalTrials.gov with the identifier NCT05976126.

Bio-Composite Nanogels Based on Chitosan and Hyaluronic Acid for the Treatment of Lung Infections
Francesca Della Sala, Marco Barretta, Mario di Gennaro, Rubina Paradiso, Giorgia Borriello, and Assunta Borzacchiello
MDPI AG
Pathogen infections constitute a serious problem in the field of lung diseases, especially in severe conditions such as chronic obstructive pulmonary disease (COPD) and acute respiratory distress syndrome (ARDS). Exacerbations of COPD and ARDS can be significantly influenced by bacterial infections from Pseudomonas aeruginosa and Staphylococcus aureus, which can hasten the decline of lung function. Moreover, the abuse of high-dose antibiotics used to treat obstinate infections is contributing to the growing issue of multidrug resistance (MDR) by microorganisms. Currently, new therapeutic strategies capable of surprising and fighting pathogens with new modalities are missing. In this framework, bio-composite nanogels (NGs) based on natural polymers with intrinsic antimicrobial properties such as chitosan (CS) and hyaluronic acid (HA) have been developed for the treatment of lung infections. The DLS and TEM results showed that NGs have a spherical shape with a size smaller than 100 nm, making it possible for them to potentially reach the lung site and evade the clearance of alveolar macrophages. FTIR spectra demonstrated that only electrostatic interactions, not chemical reactions, occur between NG precursors. Rheological analysis highlighted NGs’ injectability and mucoadhesive capacity. Moreover, an MTT assay on human lung fibroblast cells for biocompatibility evaluation showed good viability up to 48 h. Finally, an antimicrobial test on P. aeruginosa and S. aureus showed an increase in antimicrobial activity as the NG concentration increases, with a reduction in bacterial growth of around 60% at 375 μg/mL.

First description of Rhodococcus equi infection in common bottlenose dolphin (Tursiops truncatus)
Maria Oliviero, Doriana Iaccarino, Emanuele Esposito, Gianluigi Paduano, Fabio Di Nocera, Giorgia Borriello, Rubina Paradiso, Barbara degli Uberti, Maria Dimatteo, Maria Grazia Amoroso,et al.
Springer Science and Business Media LLC

Detection and quantification of Brucella abortus DNA in water buffaloes (bubalus bubalis) using droplet digital polymerase chain reaction
Giovanna Fusco, Lorena Cardillo, Ornella Valvini, Alessia Pucciarelli, Gerardo Picazio, Anna Cerrone, Michele Napoletano, Roberta Pellicanò, Maria Ottaiano, Claudio de Martinis,et al.
Informa UK Limited
Abstract Brucellosis represents a major public health concern worldwide. Human transmission is mainly due to the consumption of unpasteurized milk and dairy products of infected animals. The gold standard for the diagnosis of Brucella spp in ruminants is the bacterial isolation, but it is time-consuming. Polymerase Chain Reaction (PCR) is a quicker and more sensitive technique than bacterial culture. Droplet digital PCR (ddPCR) is a novel molecular assay showing high sensitivity in samples with low amount of DNA and lower susceptibility to amplification inhibitors. Present study aimed to develop a ddPCR protocol for the detection of Brucella abortus in buffalo tissue samples. The protocol was validated using proficiency test samples for Brucella spp by real time qPCR. Furthermore, 599 tissue samples were examined. Among reference materials, qPCR and ddPCR demonstrated same performance and were able to detect up to 225 CFU/mL. Among field samples, ddPCR showed higher sensitivity (100%), specificity and accuracy of 93.4% and 94.15%, respectively. ddPCR could be considered a promising technique to detect B. abortus in veterinary specimens, frequently characterized by low amount of bacteria, high diversity in matrices and species and poor storage conditions.

Modifications of lung microbiota structure in traumatic brain injury ventilated patients according to time and enteral feeding formulas: a prospective randomized study
A. Cotoia, R. Paradiso, G. Ferrara, G. Borriello, F. Santoro, I. Spina, L. Mirabella, K. Mariano, G. Fusco, G. Cinnella,et al.
Springer Science and Business Media LLC
Abstract Background Specialized diets enriched with immune nutrients could be an important supplement in patients (pts) with acute traumatic brain injury (TBI). Omega-3 and arginine may interact with immune response and microbiota. No data are available about the role of the specialized diets in modulating the lung microbiota, and little is known about the influence of lung microbiota structure in development of ventilator-associated pneumonia (VAP) in TBI pts. The aims of this study are to evaluate the impact of specific nutrients on the lung microbiota and the variation of lung microbiota in TBI pts developing VAP. Methods A cohort of 31 TBI pts requiring mechanical ventilation in ICU was randomized for treatment with specialized (16pts) or standard nutrition (15pts). Alpha and beta diversity of lung microbiota were analyzed from bronco Alveolar Lavage (BAL) samples collected at admission and 7 days post-ICU admission in both groups. A further analysis was carried out on the same samples retrospectively grouped in VAP or no VAP pts. Results None developed VAP in the first week. Thereafter, ten out of thirty-one pts developed VAP. The BAL microbiota on VAP group showed significant differences in beta diversity and Staphylococcus and Acinetobacter Genera were high. The specialized nutrition had influence on beta diversity that reached statistical significance only in Bray–Curtis distance. Conclusion Our data suggest that TBI patients who developed VAP during ICU stay have different structures of BAL microbiota either at admission and at 7 days post-ICU admission, while no correlation has been observed between different enteral formulas and microbiota composition in terms of richness and evenness. These findings suggest that targeting the lung microbiota may be a promising approach for preventing infections in critically ill patients.

Effect of rennet on buffalo cheese microbiota, taste and flavor
Angela M. I. Montone, Angelo Citro, Germana Colarusso, Francesca Garofalo, Rubina Paradiso, Giorgia Borriello, Domenico Vecchio, and Federico Capuano
Universidad del Zulia
Rennet is a critical element that allows the coagulation of milk caseins, the basic process of producing cheese. Traditional cheeses (Traditional Agri-food Products - PAT) are generally produced using artisanal animal rennet. The type of rennet used for cheese making, jointly with several other factors, influences the flavor and aroma of dairy products. In this study, we focused exclusively on rennet’s role in developing aromas and flavors, highlighting different tastes depending on the type of rennet used in cheesemaking. Buffalo milk was used to produce cheeses through a single cheesemaking session, where the only difference was the rennet used: commercial calf or artisanal lamb. Cheeses were stored and analyzed monthly at T0, T30, T60, and T90. Microbiota characterization (NGS technology) was performed on all samples, jointly with microbiological and sensory analysis (ISO 11035:1994). The microbiological analysis highlighted differences mainly in quantifying total bacteria count and acid lactic bacteria at T0, significantly higher in cheeses from calf rennet. Furthermore, at T60, a general reduction of all microbiological parameters in cheese based on lamb rennet was observed. The Microbiota analysis showed significant microbiological changes during the seasoning in the same product (α diversity) and between the two products obtained from industrial and artisanal rennet (β diversity). Regarding the sensory analysis, all the judges evaluated the cheeses based on lamb rennet as more appreciable due to their intense flavors and aroma. The study results confirmed the role of lamb rennet in developing flavor in buffalo cheese and, consequently, suggests a larger use of this rennet to obtain cheeses with a particular and typical taste based on buffalo milk.

First Report on Abortion Caused by Salmonella enterica subsp. enterica Serovar Enteritidis in Water Buffalo (Bubalus bubalis)
Luisa D'Angelo, Rubina Paradiso, Domenico Alfano, Marita Georgia Riccardi, Giorgia Borriello, and Giorgio Galiero
Frontiers Media SA
Salmonella enterica subsp. enterica Serovar Enteritidis is one of the major pathogens associated with enteric diseases in animals and humans. Thus, due to the importance of Salmonella spp. infections for animal production and public health, the aim of the present study was to describe the first detection of S. enteritidis in an aborted water buffalo fetus in southern Italy by characterizing the phylogroup profile and the antimicrobial susceptibility of the isolated pathogenic strains. The different clinical manifestations of salmonellosis in animals include diarrhea, abortion, pneumonia, septic arthritis, meningitis, and others, depending on the virulence of the serovars, infectious dose, and host immunity. This study reports the first case of abortion caused by Salmonella enterica subsp enterica serovar Enteritidis in water buffalo (Bubalus bubalis) in the Campania region, southern Italy. Complete necropsy was performed on the aborted water buffalo fetus under study, and samples and swabs from different organs were collected. Samples were processed by microbiological and molecular analyses to detect bacterial, viral, and protozoarian pathogens possibly responsible for abortion. Whole genome sequencing (WGS) was carried out to further characterize the isolated S. Enteritidis strain. Our findings highlight the crucial role of S. Enteritidis as a potential abortive agent in water buffalo and its presence should therefore be investigated in cases of bubaline abortion.

A type II variant of Toxoplasma gondii infects the Eurasian otter (Lutra lutra) in southern Italy
Maurizio Viscardi, Mario Santoro, Loredana Cozzolino, Giorgia Borriello, and Giovanna Fusco
Hindawi Limited
Toxoplasmosis, caused by the protozoan parasite Toxoplasma gondii, is a widespread zoonosis capable to affect a wide range of warm-blooded vertebrates. In the past two decades, T. gondii emerged as a significant aquatic pathogen with some pathogenic atypical genotypes isolated and characterized from stranded marine mammals. In contrast, no information is available for mammals in freshwater environment. Although, otters are considered highly susceptible to T. gondii infection, to date molecular evidence of T. gondii in Eurasian otter (Lutra lutra) does not exist. We report the first molecular evidence of T. gondii in a free-ranging Eurasian otter from southern Italy, and characterized the present strain as a mixed genotype type II variant, with all loci type II except PK1 (locus sequence corresponding to type II variant B), B1 (locus sequence corresponding to type II/X A), and C29-2 (locus with SNPs). Our results indicate circulation of a type II variant in freshwater environment which suggests potential risk of transmission to animals and humans. The finding of a potential pathogenic strain is of great concern for future conservation programs of the critically endangered Eurasian otter in southern Italy.

Whole-genome sequencing-based characterization of a listeria monocytogenes strain from an aborted water buffalo in southern italy
Rubina Paradiso, Marita Georgia Riccardi, Bianca Cecere, Nunzia Riccone, Roberto Scialla, Antonietta Anzalone, Anna Cerrone, Esterina De Carlo, Giorgia Borriello, and Giovanna Fusco
MDPI AG
Listeria monocytogenes is a Gram-positive pathogen causing life-threatening infections both in humans and animals. In livestock farms, it can persist for a long time and primarily causes uterine infections and encephalitis in farmed animals. Whole genome sequencing (WGS) is currently becoming the best method for molecular typing of this pathogen due to its high discriminatory power and efficiency of characterization. This study describes the WGS-based characterization of an L. monocytogenes strain from an aborted water buffalo fetus in southern Italy. The strain under study was classified as molecular serogroup IVb, phylogenetic lineage I, MLST sequence type 6, Clonal Complex 6, and cgMLST type CT3331, sublineage 6. Molecular analysis indicated the presence of 61 virulence genes and 4 antibiotic resistance genes. Phylogenetic analysis, including all the publicly available European L. monocytogenes serogroup IVb isolates, indicated that our strain clusterized with all the other CC6 strains and that different CCs were variably distributed within countries and isolation sources. This study contributes to the current understanding of the genetic diversity of L. monocytogenes from animal sources and highlights how the WGS strategy can provide insights into the pathogenic potential of this microorganism, acting as an important tool for epidemiological studies.

Long-chain polyphosphates impair SARS-CoV-2 infection and replication
Veronica Ferrucci, Dae-Young Kong, Fatemeh Asadzadeh, Laura Marrone, Angelo Boccia, Roberto Siciliano, Giuseppina Criscuolo, Camilla Anastasio, Fabrizio Quarantelli, Marika Comegna,et al.
American Association for the Advancement of Science (AAAS)
Long-chain polyphosphates inhibit SARS-CoV-2 infection by targeting a host receptor and a viral RNA polymerase.

Mycobacterium tuberculosis SIT42 Infection in an Abused Dog in Southern Italy
Lucia Vangone, Lorena Cardillo, Marita Georgia Riccardi, Giorgia Borriello, Anna Cerrone, Paolo Coppa, Roberto Scialla, Emanuela Sannino, Gianluca Miletti, Giorgio Galiero,et al.
Frontiers Media SA
A case of Mycobacterium tuberculosis infection is described in a dead adult male dog in Southern Italy. The carcass was found by the Health Authority in a gypsy encampment. It was admitted to our forensic veterinary medicine unit, with a suspicion of cruelty to the animal. Necropsy showed beating and traumatism signs, and mistreating was confirmed. Gross lesions included multiple nodular hepatic lesions, hemorrhagic enteritis with enlarged mesenteric lymph nodes, body cavity effusions, and an adrenal neoplasm. Bacteriological and molecular analyses were carried out on the liver lesions that enabled to identify M. tuberculosis SIT42 (LAM9). Drug-resistance patterns were evaluated by screening mutations on the rpoB and katG genes that showed susceptibility to both rifampin and isoniazid, respectively. Very few studies report canine tuberculosis, and little is known about the disease in Italy. To the authors' knowledge, this is the first report of Mycobacterium tuberculosis SIT42 infection in a dog in Italy.

Different non-structural carbohydrates/crude proteins (NCS/CP) ratios in diet shape the gastrointestinal microbiota of water buffalo
Rubina Paradiso, Giorgia Borriello, Sergio Bolletti Censi, Angela Salzano, Roberta Cimmino, Giorgio Galiero, Giovanna Fusco, Esterina De Carlo, and Giuseppe Campanile
MDPI AG
The microbiota of the gastrointestinal tract (GIT) are crucial for host health and production efficiency in ruminants. Its microbial composition can be influenced by several endogenous and exogenous factors. In the beef and dairy industry, the possibility to manipulate gut microbiota by diet and management can have important health and economic implications. The aims of this study were to characterize the different GIT site microbiota in water buffalo and evaluate the influence of diet on GIT microbiota in this animal species. We characterized and compared the microbiota of the rumen, large intestine and feces of water buffaloes fed two different diets with different non-structural carbohydrates/crude proteins (NSC/CP) ratios. Our results indicated that Bacteroidetes, Firmicutes and Proteobacteria were the most abundant phyla in all the GIT sites, with significant differences in microbiota composition between body sites both within and between groups. This result was particularly evident in the large intestine, where beta diversity analysis displayed clear clustering of samples depending on the diet. Moreover, we found a difference in diet digestibility linked to microbiota modification at the GIT level conditioned by NSC/CP levels. Diet strongly influences GIT microbiota and can therefore modulate specific GIT microorganisms able to affect the health status and performance efficiency of adult animals.

SERS biosensor based on engineered 2D-aperiodic nanostructure for in-situ detection of viable brucella bacterium in complex matrix
Massimo Rippa, Riccardo Castagna, Domenico Sagnelli, Ambra Vestri, Giorgia Borriello, Giovanna Fusco, Jun Zhou, and Lucia Petti
MDPI AG
Brucella is a foodborne pathogen globally affecting both the economy and healthcare. Surface Enhanced Raman Spectroscopy (SERS) nano-biosensing can be a promising strategy for its detection. We combined high-performance quasi-crystal patterned nanocavities for Raman enhancement with the use of covalently immobilized Tbilisi bacteriophages as high-performing bio-receptors. We coupled our efficient SERS nano-biosensor to a Raman system to develop an on-field phage-based bio-sensing platform capable of monitoring the target bacteria. The developed biosensor allowed us to identify Brucella abortus in milk by our portable SERS device. Upon bacterial capture from samples (104 cells), a signal related to the pathogen recognition was observed, proving the concrete applicability of our system for on-site and in-food detection.

First Detection of Listeria monocytogenes in a Buffalo Aborted Foetus in Campania Region (Southern Italy)
Claudia Esposito, Lorena Cardillo, Giorgia Borriello, Grazia Ascione, Ornella Valvini, Giorgio Galiero, and Giovanna Fusco
Frontiers Media SA
Listeria monocytogenes (LM) is the causative agent of listeriosis in both animals and humans, representing one of the most severe food-borne diseases in humans. Out of 13 serotypes, only three (i.e., 1/2a, 1/2b, and 4b) are responsible for 95% of human outbreaks of listeriosis. Ruminants have been hypothesised to represent the main natural reservoir for this pathogen and to be involved in the transmission of Listeria to humans. During pregnancy, listeriosis in ruminants cause various reproductive disorders as well as abortion. However, little is known about abortion due to LM in water buffaloes (Bubalus bubalis). In this study, we report for the first time the detection of LM in a water buffalo foetus in the region of Campania, Italy. Complete necropsy was performed, and samples and swabs from the abomasum, kidneys, liver, lungs, and spleen were collected. Microbiological and molecular analyses were carried out to detect bacterial, viral, and protozoarian abortive pathogens. The results revealed the presence of LM in the liver, lungs, and abomasum, and no other agent was detected. Isolation was confirmed by biochemical and molecular tests. Molecular serotype characterisation was performed, and serogroup IVb was identified. In conclusion, because of the zoonotic implications of our findings, this report highlights the importance of including LM in the diagnostic panel in cases of bubaline abortion.

Cerumen microbial community shifts between healthy and otitis affected dogs
Giorgia Borriello, Rubina Paradiso, Carlotta Catozzi, Roberta Brunetti, Paola Roccabianca, Marita Georgia Riccardi, Bianca Cecere, Cristina Lecchi, Giovanna Fusco, Fabrizio Ceciliani,et al.
Public Library of Science (PLoS)
Otitis externa is a common multifactorial disease in dogs, characterized by broad and complex modifications of the ear microbiota. The goal of our study was to describe the ear cerumen microbiota of healthy dogs, within the same animal and between different animals, and to compare the cerumen microbiota of otitis affected dogs with that of healthy animals. The present study included 26 healthy dogs, 16 animals affected by bilateral otitis externa and 4 animals affected by monolateral otitis externa. For each animal cerumen samples from the right and left ear were separately collected with sterile swabs, and processed for DNA extraction and PCR amplification of the 16S rRNA gene. Amplicon libraries were sequenced using an Ion Torrent Personal Genome Machine (PGM), and taxonomical assignment and clustering were performed using QIIME 2 software. Our results indicate that the bacterial community of the cerumen in healthy dogs was characterized by extensive variability, with the most abundant phyla represented by Proteobacteria, Actinobacteria, Firmicutes, Bacteroidetes and Fusobacteria. The analysis of both alpha and beta diversity between pairs of left and right ear samples from the same dog within the group of affected animals displayed higher differences than between paired samples across healthy dogs. Moreover we observed reduced bacterial richness in the affected group as compared with controls and increased variability in population structure within otitis affected animals, often associated with the proliferation of a single bacterial taxon over the others. Moreover, Staphylococcus and Pseudomonas resulted to be the bacterial genera responsible for most distances between the two groups, in association with differences in the bacterial community structure. The cerumen microbiota in healthy dogs exhibits a complex bacterial population which undergoes significant modifications in otitis affected animals.

Fatal interstitial pneumonia associated with bovine coronavirus in cows from southern italy
Maria Grazia Amoroso, Giuseppe Lucifora, Barbara Degli Uberti, Francesco Serra, Giovanna De Luca, Giorgia Borriello, Alessandro De Domenico, Sergio Brandi, Maria Concetta Cuomo, Francesca Bove,et al.
MDPI AG
An outbreak of winter dysentery, complicated by severe respiratory syndrome, occurred in January 2020 in a high production dairy cow herd located in a hilly area of the Calabria region. Of the 52 animals belonging to the farm, 5 (9.6%) died with severe respiratory distress, death occurring 3–4 days after the appearance of the respiratory signs (caught and gasping breath). Microbiological analysis revealed absence of pathogenic bacteria whilst Real-time PCR identified the presence of RNA from Bovine Coronavirus (BCoV) in several organs: lungs, small intestine (jejunum), mediastinal lymph nodes, liver and placenta. BCoV was therefore hypothesized to play a role in the lethal pulmonary infection. Like the other CoVs, BCoV is able to cause different syndromes. Its role in calf diarrhea and in mild respiratory disease is well known: we report instead the involvement of this virus in a severe and fatal respiratory disorder, with symptoms and disease evolution resembling those of Severe Acute Respiratory Syndromes (SARS).

Genotyping of Toxoplasma gondii in wild boar (Sus scrofa) in southern Italy: Epidemiological survey and associated risk for consumers
Giovanni Sgroi, Maurizio Viscardi, Mario Santoro, Giorgia Borriello, Nicola D'Alessio, Federica Boccia, Laura Pacifico, Alessandro Fioretti, Vincenzo Veneziano, and Giovanna Fusco
Wiley
AbstractToxoplasma gondii is a widespread protozoan parasite (phylum Apicomplexa), which causes a zoonotic parasitic disease, known as toxoplasmosis. The aim of this study was to evaluate the occurrence and genotypes of T. gondii in wild boars of southern Italy and thus to assess the risk of infection for consumers. The boars were inspected during the hunting season within the regional project ‘Wild Boar Emergency Plan in Campania’, and molecular analyses were performed on 338 boars analysing a total number of 884 matrices (263 brains, 310 hearts and 311 masseter muscles). Toxoplasma gondii was detected in 134 out of 338 boars (39.6%). No significant statistical difference between genders was found (χ2 = 0.15 p = .70). The prevalence was 47.1%, 39.3% and 39.2% in piglets, yearlings and adults, respectively (χ2 = 0.41; p = .81). The highest prevalence of T. gondii was found in masseter muscles (74/311, 23.8%), followed by the heart (70/310, 22.6%) and brain (58/263, 22.0%), respectively. Microsatellite (MS) analysis of 11 samples revealed eleven T. gondii genotypes (nine atypical, one belonging to type II one to type III). Most of the genotypes found were thus atypical and may be virulent in humans. Hierarchical clustering analysis showed the presence of three distinct clusters, with the majority of atypical genotypes in the GII‐GIII cluster. The high prevalence of infection in masseters highlights the potential risk for public health, considering that this muscle is commonly used to prepare raw meat products (‘guanciale’ and sausages), which may be a source of T. gondii infection in humans. Wild boars may act as an interface role between wildlife, livestock and humans. Our data highlight the urgent need to minimize the risk of infection for animals and humans by setting up a surveillance programme and preventive strategies in a One Health approach to wildlife species.

Antibiotic activity of a paraphaeosphaeria sporulosa-produced diketopiperazine against salmonella enterica
Raffaele Carrieri, Giorgia Borriello, Giulio Piccirillo, Ernesto Lahoz, Roberto Sorrentino, Michele Cermola, Sergio Bolletti Censi, Laura Grauso, Alfonso Mangoni, and Francesco Vinale
MDPI AG
A diketopiperazine has been purified from a culture filtrate of the endophytic fungus Paraphaeosphaeria sporulosa, isolated from healthy tissues of strawberry plants in a survey of microbes as sources of anti-bacterial metabolites. Its structure has been determined by nuclear magnetic resonance (NMR) and liquid chromatography–mass spectrometry (LC–MS) analyses and was found to be identical to cyclo(L-Pro-L-Phe) purified from species of other fungal genera. This secondary metabolite has been selected following bioguided-assay fractionation against two strains of Salmonella enterica, the causal agent of bovine gastroenteritis. The diketopiperazine cyclo(L-Pro-L-Phe), isolated for the first time from Paraphaeosphaeria species, showed minimum inhibitory concentration (MIC) values of 71.3 and 78.6 μg/mL against the two S. enterica strains. This finding may be significant in limiting the use of synthetic antibiotics in animal husbandry and reducing the emergence of bacterial multidrug resistance. Further in vivo experiments of P. sporulosa diketopiperazines are important for the future application of these metabolites.

Identification of Altered miRNAs in Cerumen of Dogs Affected by Otitis Externa
Cristina Lecchi, Valentina Zamarian, Giorgia Borriello, Giorgio Galiero, Guido Grilli, Mario Caniatti, Elisa Silvia D'Urso, Paola Roccabianca, Roberta Perego, Michela Minero,et al.
Frontiers Media SA
Otitis externa is one of the most common diseases in dogs. It is associated with bacteria and yeast, which are regarded as secondary causes. Cerumen is a biological substance playing an important role in the protection of ear skin. The involvement of cerumen in immune defense is poorly understood. MicroRNAs can modulate the host immune response and can provide promising biomarkers for several inflammatory and infectious disorder diagnosis. The aims of this study were to profile the cerumen miRNA signature associated with otitis externa in dogs, integrate miRNAs to their target genes related to immune functions, and investigate their potential use as biomarkers. Cerumen was collected from healthy and otitis affected dogs and the expression of miRNAs was profiled by Next Generation Sequencing; the validation of the altered miRNAs was performed using RT-qPCR. The potential ability of miRNAs to modulate immune-related genes was investigated using bioinformatics tools. The results pointed out that 32 miRNAs, of which 14 were up- and 18 down-regulated, were differentially expressed in healthy vs. otitis-affected dogs. These results were verified by RT-qPCR. To assess the diagnostic value of miRNAs, ROC analysis was carried out, highlighting that 4 miRNAs are potential biomarkers to discriminate otitis-affected dogs. Bioinformatics showed that cerumen miRNAs may be involved in the modulation of host immune response. In conclusion, we have demonstrated for the first time that miRNAs can be efficiently extracted and quantified from cerumen, that their profile changes between healthy and otitis affected dogs, and that they may serve as potential biomarkers. Further studies are necessary to confirm their diagnostic value and to investigate their interaction with immune-related genes.

Different impacts of mucr binding to the babr and virb promoters on gene expression in brucella abortus 2308
Giorgia Borriello, Veronica Russo, Rubina Paradiso, Marita Georgia Riccardi, Daniela Criscuolo, Gaetano Verde, Rosangela Marasco, Paolo Vincenzo Pedone, Giorgio Galiero, and Ilaria Baglivo
MDPI AG
The protein MucR from Brucella abortus has been described as a transcriptional regulator of many virulence genes. It is a member of the Ros/MucR family comprising proteins that control the expression of genes important for the successful interaction of α-proteobacteria with their eukaryotic hosts. Despite clear evidence of the role of MucR in repressing virulence genes, no study has been carried out so far demonstrating the direct interaction of this protein with the promoter of its target gene babR encoding a LuxR-like regulator repressing virB genes. In this study, we show for the first time the ability of MucR to bind the promoter of babR in electrophoretic mobility shift assays demonstrating a direct role of MucR in repressing this gene. Furthermore, we demonstrate that MucR can bind the virB gene promoter. Analyses by RT-qPCR showed no significant differences in the expression level of virB genes in Brucella abortus CC092 lacking MucR compared to the wild-type Brucella abortus strain, indicating that MucR binding to the virB promoter has little impact on virB gene expression in B. abortus 2308. The MucR modality to bind the two promoters analyzed supports our previous hypothesis that this is a histone-like protein never found before in Brucella.

Parasite load and strs genotyping of toxoplasma gondii isolates from mediterranean mussels (Mytilus galloprovincialis) in southern italy
Mario Santoro, Maurizio Viscardi, Federica Boccia, Giorgia Borriello, Maria Gabriella Lucibelli, Clementina Auriemma, Aniello Anastasio, Vincenzo Veneziano, Giorgio Galiero, Loredana Baldi,et al.
Frontiers Media SA
Toxoplasmosis is a zoonotic food-borne disease caused by Toxoplasma gondii, a land-derived protozoan parasite that infects a broad range of terrestrial and aquatic hosts. T. gondii may reach coastal waters via contaminated freshwater runoff and its oocysts may enter into the marine food web. Marine invertebrates as mussels being filter feeders are exposed and may concentrate T. gondii oocysts representing a potential source of infection for animals and humans. The present works investigated the prevalence, parasite burden and genotypes of T. gondii in the Mediterranean mussels (Mytilus galloprovincialis) from southern Italy. We sampled a total of 382 individual Mediterranean mussels from May to August 2018 from seven production sites in the Gulf of Naples (Campania region). An additional sample including 27 farmed Mediterranean mussels was obtained in February 2018 from a mollusk depuration plant in Corigliano Calabro (Calabria region). T. gondii DNA was detected in 43 out of 409 (10.5%) Mediterranean mussels from seven out of eight sampling sites. The number of T. gondii copies/g in the digestive gland ranged from 0.14 to 1.18. Fragment analysis of Short Tandem Repeats (STRs) at 5 microsatellite loci was performed from 10 T. gondii PCR positive samples revealing the presence of five distinct genotypes including one corresponding to type I and four atypical genotypes. These findings suggest potential implications of epidemiological importance for human and animal health because both type I and atypical genotypes could be highly pathogenic.

Screen printed based impedimetric immunosensor for rapid detection of Escherichia coli in drinking water
Martina Cimafonte, Andrea Fulgione, Rosa Gaglione, Marina Papaianni, Rosanna Capparelli, Angela Arciello, Sergio Bolletti Censi, Giorgia Borriello, Raffaele Velotta, and Bartolomeo Della Ventura
MDPI AG
The development of a simple and low cost electrochemical impedance immunosensor based on screen printed gold electrode for rapid detection of Escherichia coli in water is reported. The immunosensor is fabricated by immobilizing anti-E. coli antibodies onto a gold surface in a covalent way by the photochemical immobilization technique, a simple procedure able to bind antibodies upright onto gold surfaces. Impedance spectra are recorded in 0.01 M phosphate buffer solution (PBS) containing 10 mM Fe(CN)63−/Fe(CN)64− as redox probe. The Nyquist plots can be modelled with a modified Randles circuit, identifying the charge transfer resistance Rct as the relevant parameter after the immobilization of antibodies, the blocking with BSA and the binding of E. coli. The introduction of a standard amplification procedure leads to a significant enhancement of the impedance increase, which allows one to measure E. coli in drinking water with a limit of detection of 3 × 101 CFU mL−1 while preserving the rapidity of the method that requires only 1 h to provide a “yes/no” response. Additionally, by applying the Langmuir adsorption model, we are able to describe the change of Rct in terms of the “effective” electrode, which is modified by the detection of the analyte whose microscopic conducting properties can be quantified.

Giorgia Borriello

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