Interface interactions driven antioxidant properties in olive leaf extract/cellulose nanocrystals/poly(butylene adipate-co-terephthalate) biomaterials.

Functional packaging represents a new frontier for research on food packaging materials. In this context, adding antioxidant properties to packaging films is of interest. In this study, poly(butylene adipate-co-terephthalate) (PBAT) and olive leaf extract (OLE) have been melt-compounded to obtain novel biomaterials suitable for applications which would benefit from the antioxidant activity. The effect of cellulose nanocrystals (CNC) on the PBAT/OLE system was investigated, considering the interface interactions between PBAT/OLE and OLE/CNC. The biomaterials' physical and antioxidant properties were characterized. Morphological analysis corroborates the full miscibility between OLE and PBAT and that OLE favours CNC dispersion into the polymer matrix. Tensile tests show a stable plasticizer effect of OLE for a month in line with good interface PBAT/OLE interactions. Simulant food tests indicate a delay of OLE release from the 20 wt% OLE-based materials. Antioxidant activity tests prove the antioxidant effect of OLE depending on the released polyphenols, prolonged in the system at 20 wt% of OLE. Fluorescence spectroscopy demonstrates the nature of the non-covalent PBAT/OLE interphase interactions in π-π stacking bonds. The presence of CNC in the biomaterials leads to strong hydrogen bonding interactions between CNC and OLE, accelerating OLE released from the PBAT matrix.

Antimicrobial efficacy of Punica granatum Lythraceae peel extract against pathogens belonging to the ESKAPE group.

The improper use and abuse of antibiotics have led to an increase in multidrug-resistant (MDR) bacteria resulting in a failure of standard antibiotic therapies. To date, this phenomenon represents a leading public health threat of the 21st century which requires alternative strategies to fight infections such as the identification of new molecules active against MDR strains. In the last 20 years, natural extracts with biological activities attracted scientific interest. Following the One Health Approach, natural by-products represent a sustainable and promising alternative solution. Consistently, the aim of the present study was to evaluate the antimicrobial activity of hydro-alcoholic pomegranate peel extract (PPE) against MDR microorganisms belonging to Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. "ESKAPE" group pathogens. Through semiquantitative and quantitative methods, the PPE showed effective antimicrobial activity against Gram-positive and Gram-negative MDR bacteria. The kinetics of bactericidal action of PPE highlighted that microbial death was achieved in a time- and dose-dependent manner. High concentrations of PPE exhibited antioxidant activity, providing a protective effect on cellular systems and red blood cell membranes. Finally, we report, for the first time, a significant intracellular antibacterial property of PPE as highlighted by its bactericidal action against the staphylococcal reference strain and its bacteriostatic effect against clinical resistant strain in the HeLa cell line. In conclusion, due to its characterized content of polyphenolic compounds and antioxidant activity strength, the PPE could be considered as a therapeutic agent alone or in conjunction with standard antibiotics against challenging infections caused by ESKAPE pathogens.

Eco-Friendly Sanitization of Indoor Environments: Effectiveness of Thyme Essential Oil in Controlling Bioaerosol Levels and Disinfecting Surfaces.

Bioaerosols and pathogens in indoor workplaces and residential environments are the primary culprits of several infections. Techniques for sanitizing air and surfaces typically involve the use of UV rays or chemical sanitizers, which may release chemical residues harmful to human health. Essential oils, natural substances derived from plants, which exhibit broad antimicrobial properties, could be a viable alternative for air and surface sanitation. The objective of this study has been to investigate the efficacy of thyme essential oil (TEO) in environmental sanitation processes. In Vitro assays through agar well diffusion, disk volatilization and tube dilution methods revealed significant antimicrobial activity of TEO 100% against foodborne and environmental isolates, with both bacteriostatic/fungistatic and bactericidal/fungicidal effects. Therefore, aqueous solutions of TEO 2.5% and 5% were formulated for air sanitation through nebulization and surface disinfection via direct contact. Bioaerosol samples and surface swabs were analyzed before and after sanitation, demonstrating the efficacy of aqueous solutions of TEO in reducing mesophilic and psychrophilic bacteria and environmental fungi levels in both air and on surfaces. The obtained results prove the antimicrobial potential of aqueous solutions of TEO in improving indoor air quality and surface cleanliness, suggesting thyme essential oil as an effective and safe natural sanitizer with minimal environmental impact compared to dangerous chemical disinfectants.

Vine-Winery Byproducts as Precious Resource of Natural Antimicrobials: In Vitro Antibacterial and Antibiofilm Activity of Grape Pomace Extracts against Foodborne Pathogens.

Grape pomace is the main by-product of vine-winery chains. It requires adequate treatment and disposal but is also an economically underused source of bioactive plant secondary metabolites. This study aimed to investigate the antibacterial effects of polyphenolic extracts from Aglianico (Vitis vinifera L.) grape pomace. In particular, hydroethanolic extracts obtained via an ultrasonic-assisted extraction technique were selected for antimicrobial tests. The extracts were screened for their antibacterial effects against foodborne pathogens that were both Gram-positive, in the case of Staphylococcus aureus and Bacillus cereus, and Gram-negative, in the case of Escherichia coli and Salmonella enterica subsp. enterica serovar Typhimurium, showing variable bacteriostatic and bactericidal effects. In addition, our results demonstrated that the tested grape pomace extracts can reduce the inhibitory concentration of standard antibiotics. Interestingly, selected extracts inhibited biofilm development by S. aureus and B. cereus. Overall, these new insights into the antibacterial properties of grape pomace extracts may represent a relevant step in the design of novel therapeutic tools to tackle foodborne diseases, and in the management of resistant biofilm-related infections.

Antibacterial and Antibiofilm Efficacy of Thyme (Thymus vulgaris L.) Essential Oil against Foodborne Illness Pathogens, Salmonella enterica subsp. enterica Serovar Typhimurium and Bacillus cereus.

Nowadays, the wide spread of foodborne illness and the growing concerns about the use of synthetic food additives have shifted the focus of researchers towards essential oils (EOs) as possible antimicrobials and preservatives of natural origin. Thanks to their antimicrobial properties against pathogenic and food spoilage microorganisms, EOs have shown good potential for use as alternative food additives, also to counteract biofilm-forming bacterial strains, the spread of which is considered to be among the main causes of the increase in foodborne illness outbreaks. In this context, the aim of this study has been to define the antibacterial and antibiofilm profile of thyme (Thymus vulgaris L.) essential oil (TEO) against widespread foodborne pathogens, Salmonella enterica subsp. enterica serovar Typhimurium and Bacillus cereus. TEO chemical composition was analyzed through gas chromatography-mass spectrometry (GC-MS). Preliminary in vitro antibacterial tests allowed to qualitatively verify TEO efficacy against the tested foodborne pathogens. The subsequent determination of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) values allowed to quantitatively define the bacteriostatic and bactericidal effects of TEO. To evaluate the ability of essential oils to inhibit biofilm formation, a microplate assay was performed for the bacterial biofilm biomass measurement. Results suggest that TEO, rich in bioactive compounds, is able to inhibit the growth of tested foodborne bacteria. In addition, the highlighted in vitro anti-biofilm properties of TEO suggest the use of this natural agent as a promising food preservative to counteract biofilm-related infections in the food industry.

Towards Green Strategies of Food Security: Antibacterial Synergy of Essential Oils from Thymus vulgaris and Syzygium aromaticum to Inhibit Escherichia coli and Staphylococcus aureus Pathogenic Food Isolates.

Foodborne diseases continue to represent an important public health issue. The control of food spoilage and pathogenic microorganisms is achieved mainly by synthetic chemicals, unfortunately associated to several undesirable aspects. The growing requirement for new and safe alternative strategies has resulted in the research of agents from natural sources with antimicrobial properties, such as essential oils (EOs). This study's purpose was to define the antibacterial profile of thyme (Thymus vulgaris) and cloves (Syzygium aromaticum) essential oils against both Gram-positive and Gram-negative important foodborne pathogenic bacteria. Gas chromatography mass spectrometry analysis was performed for EOs' chemical composition. Qualitative in vitro antimicrobial assays (i.e., agar well diffusion method and disk-volatilization method) allowed for verification of the efficacy of EOs, used individually and in binary combination and both in liquid and vapor phase, against Staphylococcus aureus and Escherichia coli food isolates. Minimal inhibitory concentrations and minimal bactericidal concentration values have been used to quantitatively measure the antibacterial activity of EOs, while the fractional inhibitory concentration index has been considered as a predictor of in vitro antibacterial synergistic effects. The microbiological tests suggest that thyme and cloves EOs, rich in bioactive compounds, are able to inhibit the growth of tested foodborne bacteria, especially in vapor phase, also with synergistic effects. Results provide evidence to consider the tested essential oils as promising sources for development of new, broad-spectrum, green food preservatives.

Molecular Epidemiology of Genital Infections in Campania Region: A Retrospective Study.

This study provides updated information on the prevalence and co-infections caused by genital microorganisms and pathogens: Mycoplasma genitalium, Mycoplasma hominis, Ureaplasma parvum, Ureaplasma urealyticum, Trichomonas vaginalis, and Gardnerella vaginalis, by retrospectively analyzing a cohort of patients living in the Naples metropolitan area, Campania region, Southern Italy. To investigate the genital infections prevalence in clinical specimens (vaginal/endocervical swabs and urines) collected from infertile asymptomatic women and men from November 2018 to December 2020, we used a multiplex real-time PCR assay. Of the 717 specimens collected, 302 (42.1%) resulted positive for at least one of the targets named above. Statistically significant differences in genital prevalence of selected microorganisms were detected in both women (62.91%) and men (37.08%). G. vaginalis and U. parvum represented the most common findings with an 80.2% and 16.9% prevalence in vaginal/endocervical swabs and first-voided urines, respectively. Prevalence of multiple infections was 18.18% and 8.19% in women and men, respectively. The most frequent association detected was the co-infection of G. vaginalis and U. parvum with 60% prevalence. Our epidemiological analysis suggests different infection patterns between genders, highlighting the need to implement a preventative screening strategy of genital infections to reduce the complications on reproductive organs.

Chestnut Shell Tannins: Effects on Intestinal Inflammation and Dysbiosis in Zebrafish.

The aim of the present study was to test the possible ameliorative efficacy of phytochemicals such as tannins on intestinal inflammation and dysbiosis. The effect of a chestnut shell (Castanea sativa) extract (CSE) rich in polyphenols, mainly represented by tannins, on k-carrageenan-induced intestinal inflammation in adult zebrafish (Danio rerio) was tested in a feeding trial. Intestinal inflammation was induced by 0.1% k-carrageenan added to the diet for 10 days. CSE was administered for 10 days after k-carrageenan induced inflammation. The intestinal morphology and histopathology, cytokine expression, and microbiota were analyzed. The k-carrageenan treatment led to gut lumen expansion, reduction of intestinal folds, and increase of the goblet cells number, accompanied by the upregulation of pro-inflammatory factors (TNFα, COX2) and alteration in the number and ratio of taxonomic groups of bacteria. CSE counteracted the inflammatory status enhancing the growth of health helpful bacteria (Enterobacteriaceae and Pseudomonas), decreasing the pro-inflammatory factors, and activating the anti-inflammatory cytokine IL-10. In conclusion, CSE acted as a prebiotic on zebrafish gut microbiota, sustaining the use of tannins as food additives to ameliorate the intestinal inflammation. Our results may be relevant for both aquaculture and medical clinic fields.

In silico analysis of the antimicrobial activity of phytochemicals: towards a technological breakthrough.

BACKGROUND: The complications associated with infections from pathogens increasingly resistant to traditional drugs lead to a constant increase in the mortality rate among those affected. In such cases the fundamental purpose of the microbiology laboratory is to determine the sensitivity profile of pathogens to antimicrobial agents. This is an intense and complex work often not facilitated by the test's characteristics. Despite the evolution of the Antimicrobial Susceptibility Testing (AST) technologies, the technological breakthrough that could guide and facilitate the search for new antimicrobial agents is still missing. METHODS: In this work, we propose the experimental use of in silico instruments, particularly feedforward Multi-Layer Perceptron (MLP) Artificial Neural Network, and Genetic Programming (GP), to verify, but also to predict, the effectiveness of natural and experimental mixtures of polyphenols against several microbial strains. RESULTS: We value the results in predicting the antimicrobial sensitivity profile from the mixture data. Trained MLP shows very high correlations coefficients (0,93 and 0,97) and mean absolute errors (110,70 and 56,60) in determining the Minimum Inhibitory Concentration and Minimum Microbicidal Concentration, respectively, while GP not only evidences very high correlation coefficients (0,89 and 0,96) and low mean absolute errors (6,99 and 5,60) in the same tasks, but also gives an explicit representation of the acquired knowledge about the polyphenol mixtures. CONCLUSIONS: In silico tools can help to predict phytobiotics antimicrobial efficacy, providing an useful strategy to innovate and speed up the extant classic microbiological techniques.

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

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 in vitro 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 Streptococcus mutans, Streptococcus oralis, Streptococcus mitis, and Rothia dentocariosa oral isolates, suggesting that the new natural sources are rich in promising compounds able to counteract biofilm-related infections.

In vitro Synergy of Polyphenolic Extracts From Honey, Myrtle and Pomegranate Against Oral Pathogens, S. mutans and R. dentocariosa.

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 Streptococcus mutans and Rothia dentocariosa. 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 in vitro. 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 in vitro 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.

Inducing Meningococcal Meningitis Serogroup C in Mice via Intracisternal Delivery.

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.

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

In serogroup C Neisseria meningitidis, the cssA (siaA) gene codes for an UDP-N-acetylglucosamine 2-epimerase that catalyzes the conversion of UDP-N-acetyl-α-d-glucosamine into N-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 cssA 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 (LD50) of the wild-type strain 93/4286 was about four orders of magnitude lower than that of the cssA 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 corpus callosum, indicating high tropism of meningococci exposing sialic acids toward this brain structure and a specific involvement of the corpus callosum in the mouse model of meningococcal meningitis.

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

AIM: To evaluate the antimicrobial activity of hydroalcoholic extracts of pomegranate (Punica granatum 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 Streptococcus mutans Clarke ATCC® 25175™ strain and Rothia dentocariosa clinical isolate. RESULTS: Peel extracts inhibit effectively the growth and survival of S. mutans ATCC 25175 strain and R. dentocariosa clinical isolate with MIC and MBC values of 10 µg/µl and 15 µg/µl, respectively. Furthermore, the pomegranate juice extract showed high inhibitory activity against S. mutans ATCC 25175 strain with a MIC value of 25 µg/µl and a MBC value of 40 µg/µl, whereas, against R. dentocariosa, it has displayed a moderate inhibitory activity, with MIC and MBC values of 20 µg/µl and 140 µg/µl, respectively. CONCLUSIONS: In vitro 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.

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

Novel Approach for Evaluation of Bacteroides fragilis Protective Role against Bartonella henselae Liver Damage in Immunocompromised Murine Model.

Bartonella henselae is a gram-negative facultative intracellular bacterium and is the causative agent of cat-scratch disease. Our previous data have established that Bacteroides fragilis colonization is able to prevent B. henselae 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 B. henselae in immunocompromised hosts, SCID mice were co-infected with B. fragilis wild type or its mutant B. fragilis 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 B. henselae and co-infected with B. henselae/B. fragilis ΔPSA presented the major echostructural alterations. Half of the mice infected with B. henselae and all those co-infected with B. henselae/B. fragilis ΔPSA have showed an altered hepatic echogenicity compared with the renal cortex. The echogenicity score of co-infected mice with B. henselae/B. fragilis 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 B. fragilis wild type while those infected with B. fragilis ΔPSA showed the presence of collagen around the main vessels compared with the PBS control group. The liver samples of mice infected with B. henselae showed macro-areas rich in collagen, stellate cells, and histiocytic cells. Interestingly, our data demonstrated that immunocompromised SCID mice infected with B. henselae and co-infected with B. henselae/B. fragilis 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.

Inhibitory effect of pomegranate (Punica granatum L.) polyphenol extracts on the bacterial growth and survival of clinical isolates of pathogenic Staphylococcus aureus and Escherichia coli.

In the present study major polyphenols of pomegranate arils and peel by-products were extracted in 50% (v/v) aqueous ethanol, characterized and used in microbiological assays in order to test antimicrobial activity against clinically isolated human pathogenic microorganisms. Total concentration of polyphenols and in vitro antioxidant properties were determined by the Folin-Ciocalteu and DPPH methods, respectively. The most abundant bioactive molecules, including anthocyanins, catechins, tannins, gallic and ellagic acids were identified by RP-HPLC-DAD, also coupled to off-line matrix assisted laser desorption/ionization (MALDI-TOF) mass spectrometry (MS). The inhibitory spectrum of extracts against test microorganisms was assessed by the agar well-diffusion method. Data herein indicated that both pomegranate aril and peel extracts have an effective antimicrobial activity, as evidenced by the inhibitory effect on the bacterial growth of two important human pathogens, including Staphylococcus aureus and Escherichia coli, which are often involved in foodborne illness.

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

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.

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

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 ß 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.

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