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One of the most-used food contact materials is stainless steel (AISI 304L or AISI 316L), owing to its high mechanical strength, cleanability, and corrosion resistance. However, due to the presence of minimal crevices, stainless-steel is subject to microbial contamination with consequent significant reverb on health and industry costs due to the lack of effective reliability of sanitizing agents and procedures. In this study, we evaluated the noncytotoxic effect of an amorphous SiO x C y H z coating deposited on stainless-steel disks and performed a time-course evaluation for four Gram-negative bacteria and four Gram-positive bacteria. A low cytotoxicity of the SiO x C y H z coating was observed; moreover, except for some samples, a five-logarithm decrease was visible after 1 h on coated surfaces without any sanitizing treatment and inoculated with Gram-negative and Gram-positive bacteria. Conversely, a complete bacterial removal was observed after 30 s -1 min application of alcohol and already after 15 s under UVC irradiation against both bacterial groups. Moreover, coating deposition changed the wetting behaviors of treated samples, with contact angles increasing from 90.25° to 113.73°, realizing a transformation from hydrophilicity to hydrophobicity, with tremendous repercussions in various technological applications, including the food industry.

The emergence of antibiotic-resistant bacteria has become a major concern worldwide. This trend indicates the need for alternative agents to antibiotics, such as natural compounds of plant origin. Using agar disc diffusion and minimum inhibitory concentration (MIC) assays, we investigated the antimicrobial activity of Citrus aurantium (AEO), Citrus x limon (LEO), Eucalyptus globulus (EEO), Melaleuca alternifolia (TTO), and Cupressus sempervirens (CEO) essential oils (EOs) against three representatives of antibiotic-resistant pathogens and respective biofilms: vancomycin-resistant enterococci (VRE), methicillin-resistant Staphylococcus aureus (MRSA), and extended-spectrum β-lactamase (ESBL)-producing Escherichia coli . Using the checkerboard method, the efficacy of the EOs alone, in an association with each other, or in combination with the reference antibiotics was quantified by calculating fractional inhibitory concentrations (FICs). All the EOs displayed antibacterial activity against all strains to different extents, and TTO was the most effective. The results of the EO-EO associations and EO-antibiotic combinations clearly showed a synergistic outcome in most tests. Lastly, the effectiveness of EOs both alone and in association or combination against biofilm formed by the antibiotic-resistant strains was comparable to, and sometimes better than, that of the reference antibiotics. In conclusion, the combination of EOs and antibiotics represents a promising therapeutic strategy against antibiotic-resistant bacteria, even protected inside biofilms, which can allow decreasing the concentrations of antibiotics used.

Stainless steel, widely present in the food industry, is frequently exposed to bacterial colonization with possible consequences on consumers' health. 288 stainless steel disks with different roughness (0.25, 0.5 and 1 μm) were challenged with four Gram-negative ( Escherichia coli ATCC 25922, Salmonella typhimurium ATCC 1402, Yersinia enterocolitica ATCC 9610 and Pseudomonas aeruginosa ATCC 27588) and four Gram-positive bacteria ( Staphylococcus aureus ATCC 6538, Enterococcus faecalis ATCC 29212, Bacillus cereus ATCC 14579 and Listeria monocytogenes NCTT 10888) and underwent three different sanitizing treatments (UVC, alcohol 70% v/v and Gold lotion). Moreover, the same procedure was carried out onto the same surfaces after a nanotechnological surface coating (nanoXHAM ® D). A significant bactericidal effect was exerted by all of the sanitizing treatments against all bacterial strains regardless of roughness and surface coating. The nanoXHAM ® D coating itself induced an overall bactericidal effect as well as in synergy with all sanitizing treatments regardless of roughness. Stainless steel surface roughness is poorly correlated with bacterial adhesion and only sanitizing treatments can exert significant bactericidal effects. Most of sanitizing treatments are toxic and corrosive causing the onset of crevices that are able to facilitate bacterial nesting and growth. This nanotechnological coating can reduce surface adhesion with consequent reduction of bacterial adhesion, nesting, and growth.

One of the main concerns of the food industry is microbial adhesion to food contact surfaces and consequent contamination. We evaluated the potential bacteriostatic/bactericidal efficacy of aluminum surfaces with different large-scale roughness (0.25, 0.5 and 1 μm) before and after the surface treatment with a special anodizing based on titanium oxide nanotechnology (DURALTI ® ) and after 3 different sanitizing treatments, e.g., UV, alcohol and a natural product named Gold lotion. Four Gram-negative ( Escherichia coli ATCC 25922, Salmonella typhimurium ATCC 1402, Yersinia enterocolitica ATCC 9610 and Pseudomonas aeruginosa ATCC 27588) and four Gram-positive ( Staphylococcus aureus ATCC 6538, Enterococcus faecalis ATCC 29212, Bacillus cereus ATCC 14579 and Listeria monocytogenes NCTT 10888) bacteria were screened. As far as concerns aluminum surfaces without nanotechnology surface treatment, an overall bacteriostatic effect was observed for all strains with respect to the initial inoculum that was 10 6 CFU/mL. Conversely, an overall bactericidal effect was observed both for Gram-negative and -positive bacteria on DURALTI ® -treated aluminum disks, regardless of roughness and sanitizing treatment. These results are innovative in terms of the great potential of the antibacterial activity of nanotechnologically treated food contact surfaces and their combination with some sanitizing agents that might be exploited in the food industry.

The antilisterial activity of Thymus vulgaris, Salvia officinalis essential oils (EOs) and bacteriocin bacLP17 (previously isolated from seafood) was determined, using the compounds alone and in combination. The Disk Diffusion, Minimal Inhibitory Concentration (MIC) and Agar Well Diffusion assays were used to evaluate the effectiveness of the compounds against 12 Listeria monocytogenes in planktonic form, whereas the anti-Listeria biofilm activity was determined against the same strains in optical density (O.D.) at 570 nm, with crystal violet staining method. The lowest MIC values resulted for T. vulgaris EO and bacLp17 (0.5 μl/ml and 2 μl/ml, respectively). The combinations with the best results, expressed as FIC-Index, were T. vulgaris/S. officinalis EOs and EOs/bacLp17. The anti-biofilm activity of single EOs and bacLP17 was similar, whereas the combined use of the two kinds of EOs led to a synergistic activity. Lastly, the best anti-biofilm effect was observed with the combination bacLP17/S. officinalis and bacLP17/T. vulgaris, compared to both control and the single use of the EOs. The present study suggests that the combination of natural compounds such as T. vulgaris, S. officinalis EOs and bacLp17 may be a useful approach to the control of planktonic and sessile cells of L. monocytogenes in seafood products., Competing Interests: Declaration of Competing Interest The author reports no conflicts of interest in this work., (Copyright © 2020 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Sixty-nine Lactic Acid Bacteria (LAB) and bifidobacteria were isolated and identified from Italian dairy products (raw milk, cream, butter, soft cheese and yoghurt) to find new antimicrobial compounds to use as food bio-preservatives. All the isolates were preliminarily screened by the deferred antagonism method for bacteriocin production. Afterwards, to evaluate the release of bacteriocin in liquid medium, the Cell-Free Supernatant Fluid (CFSF) of the best producers was tested by agar well diffusion assay. The study allowed the selection of three bacteriocin producing strains ( Enterococcus faecium E23, Bifidobacterium thermophilum B23 and Lactobacillus bulgaricus L21), endowed with the strongest and broadest inhibitory capability against the pathogen Listeria monocytogenes . The molecular characteristics and the chemical-physical properties of both producers and the respective bacteriocins were studied and compared. The results showed that E. faecium E23 was the best producer strain and its class IIa bacteriocins, called enterocin E23, exhibited a good spectrum of activity towards L. monocytogenes . Enterocin E23 was stable over a wide range of pH and at low temperatures for at least four months and, for this reason, it can be employed in refrigerated foods for the control of L. monocytogenes , the major concern in dairy products.

The antimicrobial activity of different essential oils (EOs) from the Lamiaceae family was evaluated on Streptococcus agalactiae , Candida albicans , and lactobacilli. S. agalactiae is the main cause of severe neonatal infections, such as sepsis, meningitis, and pneumonia. C. albicans is a primary causative agent of vulvovaginal candidiasis, a multifactorial infectious disease of the lower female reproductive tract. Lactobacilli represent the dominant bacterial species of the vaginal flora and constitute the natural defense against pathogens. On the basis of the preliminary results, the attention was focused on the EOs from Lavandula x intermedia Emeric ex Loisel. and Mentha arvensis L. By using gas ghromatography (GS) retention data and mass spectra, it was possible to identify more than 90% of the total composition of the EO samples. The minimal inhibitory concentration (MIC) and anti-biofilm activity of the two EOs were determined against all isolated strains, using the EOs by themselves or in combination with each other and with drugs (erythromycin and fluconazole). The results showed a good antimicrobial and anti-biofilm activity of both EOs and a synergistic effect, leading to the best results against all the strains, resulted using the combinations EOs/EOs and antimicrobials/EOs.

Pseudomonas aeruginosa is a frequent causative agent of healthcare-associated diseases, but recently, other members of the Pseudomonas genus have been recognized to cause human colonization and infection. Since the aquatic environment could be an important source of contamination, we studied the drug resistance and virulence profiles in Pseudomonas species isolated from healthcare water systems. 17 Pseudomonas spp. out of 57 were randomly selected and their drug resistance and virulence profiles were later evaluated. Based on the positivity to the tests, the adhesion capability and biofilm formation on polystyrene and glass surfaces were studied in 6 strains, each belonging to different species. Six Pseudomonas strains (35%) were α-hemolytic, nine (53%) showed a positivity to the gelatinase test, and P. acidovorans 2R only was capable to degrade DNA. All Pseudomonas strains presented urease activity and the production of siderophores was widely observed (64,7%). Most of the strains showed one of the three types of motilities, 15 Pseudomonas (88.23%) resulted bacteriocin producers and all strains were resistant to one or more antibiotics. Lastly, among the six selected strains, P. aeruginosa 98.5 and P. fluorescens 97.4 were the best biofilm producers. Our study has highlighted how the majority of isolates shows biological characteristics that contribute to the pathogenicity of Pseudomonas. These features emphasize the virulence potentiality of other members of the Pseudomonas genus besides Pseudomonas aeruginosa, making them potentially pathogenic, especially against immunocompromised individuals.

The aim of this study was to analyze the antibacterial activity of four essential oils (EOs), Melaleuca alternifolia , Eucalyptus globulus , Mentha piperita, and Thymus vulgaris, in preventing the development and spread of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae , metallo-beta-lactamase (MBL)-producing Pseudomonas aeruginosa and carbapenemase (KPC)-producing Klebsiella pneumoniae . A total of 60 strains were obtained from the stock collection from the Microbiology Laboratory of Hesperia Hospital, Modena, Italy. Twenty ESBL-producing E. coli , 5 K. pneumoniae , 13 KPC-producing K. pneumoniae, and 20 MBL-producing P. aeruginosa were cultured and reconfirmed as ESBL and carbapenamase producers. Polymerase chain reaction was used for the detection of genes responsible for antibiotic resistance (ESBL and KPC/MBL). Antibacterial activity of the EOs was determined using the agar disk diffusion assay, and minimal inhibitory concentrations (MICs) were also evaluated. Lastly, adhesion capability and biofilm formation on polystyrene and glass surfaces were studied in 24 randomly selected strains. M. alternifolia and T. vulgaris EOs showed the best antibacterial activity against all tested strains and, as revealed by agar disk diffusion assay, M. alternifolia was the most effective, even at low concentrations. This effect was also confirmed by MICs, with values ranging from 0.5 to 16 µg/mL and from 1 to 16 µg/mL, for M. alternifolia and T. vulgaris EOs, respectively. The EOs' antibacterial activity compared to antibiotics confirmed M. alternifolia EO as the best antibacterial agent. T. vulgaris EO also showed a good antibacterial activity with MICs lower than both reference antibiotics. Lastly, a significant anti-biofilm activity was observed for the two EOs ( *P < 0.05 and **P < 0.01 for M. alternifolia and T. vulgaris EOs, respectively). A good antibacterial and anti-biofilm activity of M. alternifolia and T . vulgari s EOs against all selected strains was observed, thus demonstrating a future possible use of these EOs to treat infections caused by ESBL/carbapenemase-producing strains, even in association with antibiotics.

Schinus molle (L.) is a dioecious plant of the Anacardiaceae family, originating in South America and currently widespread in many regions throughout the world. In this work leaf extracts and derived low-pressure column chromatography (LPCC) fractions of S. molle L. male and female plants were investigated for the antimicrobial activity. Leaf extracts were tested on microbes Escherichia coli , Pseudomonas aeruginosa , Staphylococcus aureus , Enterococcus faecalis , Candida albicans and Bacillus subtilis . Furthermore, the extracts showing antimicrobial activity were fractionated by LPCC and the obtained fractions tested on the same microorganism strains. Positive fractions were investigated by gas-chromatography/mass spectrometry (GC-MS) and were seen to be rich in sesquiterpenes, sesquiterpenoids and other terpens. The obtained effects highlighted the antimicrobial properties of S. molle (L.) leaf compounds and revealed their importance as a source of bioactive molecules of potential pharmaceutical interest. To our knowledge, this is the first paper reporting investigations on the chemical composition of the extracts and derived positive fractions from Schinus molle (L.) plants grown in central Italy.

Despite its long half-life and physiological role, elastin undergoes irreversible changes (i.e elastolysis and/or calcification) impairing resilience of soft connective tissues. At present, it is still undefined: 1) to which extent elastin fibers have to be fragmented in order to increase their susceptibility to calcify; 2) which is the contribution of ionic environment on elastin mineralization; 3) why, in the same tissue area, mineralized coexist with non-mineralized fibers. The in vitro mineralization process was investigated on insoluble elastin, hydrolyzed or not-hydrolyzed, and incubated in different cell-free ionic environments. Mineral deposition is favored on hydrolyzed fibrillar structures due to exposure of multiple charged sites increasing the adsorption of Ca 2+ that can attract phosphate and increase the local ion concentration over the point of supersaturation, representing the minimum requirement for hydroxyapatite nucleation sites. At physiological pH, the degree of elastin mineralization is influenced by hydrolysis and complexity of medium composition, since ionic species, as sodium, potassium, magnesium, in addition to calcium and phosphorus, interfere with the calcification process. These findings broaden the knowledge on the factors controlling hydroxyapatite deposition on insoluble elastin and can also explain why, in vivo, calcified and non-calcified fibers can be observed within the same tissue., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: We investigated the virulence factors, genes, antibiotic resistance patterns, and genotypes (VRE and ESBL/AmpC) production in Enterococci and Enterobacteriaceae strains isolated from fecal samples of humans, dogs, and cats., Methods: A total of 100 fecal samples from 50 humans, 25 dogs, and 25 cats were used in the study. MICs of nine antimicrobials were determined using the broth microdilution method. Polymerase chain reaction was used for the detection of genes responsible for antibiotic resistance (VRE and ESBL/AmpC) and virulence genes both in Enterococcus species, such as cytolysin (cylA, cylB, cylM), aggregation substance (agg), gelatinase (gelE), enterococcal surface protein (esp), cell wall adhesins (efaAfs and efaAfm), and in Enterobacteriaceae , such as cytolysin (hemolysin) and gelatinase production (afa, cdt, cnf1, hlyA, iutA, papC, sfa)., Results: Enterococcus faecium was the most prevalent species in humans and cats, whereas Enterococcus faecalis was the species isolated in the remaining samples. A total of 200 Enterobacteriaceae strains were also detected, mainly from humans, and Escherichia coli was the most frequently isolated species in all types of samples. In the Enterococcus spp, the highest percentages of resistance for ampicillin, amoxicillin/clavulanate, erythromycin, tetracycline, ciprofloxacin, teicoplanin, and vancomycin were detected in cat isolates (41.6%, 52.8%, 38.9%, 23.6%, 62.5%, 20.8%, and 23.6% respectively), and in E. co li , a higher rate of resistance to cefotaxime and ceftazidime emerged in cat and dog samples, if compared with humans (75.4% and 66.0%, 80.0% and 71.4%, and 32.0% and 27.2%, respectively). Regarding the total number of enterococci, 5% and 3.4% of the strains were vancomycin and teicoplanin resistant, and the vancomycin resistance (van A) gene has been detected in all samples by PCR amplification. All the Enterobacteriaceae strains were confirmed as ESBL producers by PCR and sequencing, and the most frequent ESBL genes in E. coli strains from humans and pet samples were bla CTX-M-1 and bla CTX-M-15 ., Conclusions: Our results provide evidence that one or more virulence factors were present in both genera, underlining again the ability of pet strains to act as pathogens., Competing Interests: The authors declare no conflict of interest.

The antibacterial activity of Pimpinella anisum L., Cinnamomum zeylanicum , Syzygium aromaticum , and Cuminum cyminum L. essential oils (EOs) against some common pathogenic microorganisms ( Staphylococcus aureus ATCC 6538, Staphylococcus epidermidis ATCC 14990, Enterococcus faecalis ATCC 29212, Streptococcus pyogenes ATCC 1915, Escherichia coli ATCC 8739, Pseudomonas aeruginosa ATCC 27853, Aeromonas hydrophila ATCC 7966, Proteus mirabilis ATCC 10005, Klebsiella pneumoniae ATCC 13883, and Candida albicans ATCC 10231) and their biofilms was studied. The EOs inhibitory effects were evaluated by both Agar Well Diffusion assay and Minimum Inhibitory Concentration (MIC) determination. The most active EOs, cinnamon and cloves, were also tested on 18, 24, 48, 72 hours mature biofilms. Cinnamon and cloves exhibited the best results showing a significant activity against all the tested bacteria. Concerning biofilm, results suggest that Cinnamomum zeylanicum oil may be a useful approach to impair the biofilm produced by the tested Gram-negative bacteria. [Formula: see text].

We studied the antibacterial and anti-biofilm properties of MEDSTER 2000, a pH neutral biodegradable mixed acidic peroxide disinfectant belonging to the class IIb medical device which has been designed for decontamination and cold sterilization of hospital instruments. The broth microdilution method was used to define the antibacterial activity against planktonic form of both classified bacteria and antibiotic resistant strains of clinical source, whereas effectiveness toward their biofilm was determined on mature biofilm, grown both on plastic and stainless steel surfaces. The results showed that for the planktonic form the antibacterial activity of MEDSTER 2000 was already observed after 10 min at the lowest concentration (0.1%), and this effect was not exposure-and/or concentration-dependent. After the same time of exposure at the concentration of 2% the disinfectant was able to completely eradicate all tested bacteria grown in sessile form on both surfaces, with a greater than 6 log CFU/cm 2 reduction in viable cells. This result is supported by the microscope observation by crystal violet and live/dead assays. For the high antibacterial and anti-biofilm ability emerged, MEDSTER 2000 could represent a new and more effective approach for semicritical devices that need a high-level disinfection and could not sustain the process of heat sterilization.

The antibacterial activity of Rosmarinus officinalis L. and Thymus vulgaris L. essential oils (EOs), and their combination against food-borne and spoilage bacteria ( Listeria monocytogenes , Salmonella enteritidis , Yersinia enterocolitica , Escherichia coli and Pseudomonas spp.) was determined. The EOs inhibitory effect was evaluated both in vitro by using the disk diffusion assay and the minimum inhibitory concentration (MIC) determination, and on food by using an artificially contaminated ready-to-eat (RTE) vegetables. The results showed that the lowest MIC values were obtained with R. officinalis and T. vulgaris EOs against E. coli (4 and 8 μL/mL, respectively). The incorporation of the EOs alone or their combination in RTE vegetables reduced the viable counts of all the tested strains. Lastly, in the on food study we simulated the worst hygienic conditions, obtaining results that can be considered a warranty of safety.

Methodology . Biochemical and molecular methods were used to identify 100 lactobacilli isolated from rectal swabs. Among these, L. paracasei ssp. paracasei LP5 and L. brevis LP9 showed significant antibacterial activity against S. agalactiae and L. monocytogenes . Accordingly, characterization of their bacteriocins, BacLP5 and BacLP9, was conducted to obtain information on their kinetic production, sensitivity to chemico-physical parameters and molecular weight. To investigate the possible use of the two Lactobacillus strains as probiotics, their gastrointestinal resistance, cellular adhesiveness and sensitivity to antibiotics were also studied. Results . The obtained data show that BacLP5 and BacLP9 most likely belong to class II bacteriocins and both have a molecular weight of approximately 3 kDa. The production of BacLP5 and BacLP9 started after 4 h (40 and 80 AU ml -1 ), respectively. Both of the Lactobacillus strains survived gastric and intestinal juices well and showed adhesive capability on HEp-2 cells. Conclusion . Due to their peculiar antimicrobial characteristics, L. paracasei ssp. paracasei LP5 and L. brevis LP9 are suitable for use in the treatment of vaginal disorders, through both oral and transvaginal administration.

This study addressed the bacteriocin production in 116 lactic acid bacteria isolated from 143 fish and seafood samples. The screening for the production of antibacterial substances allowed for the selection of 16 LAB isolates endowed with inhibitory capability. Bacteriocins (bacLP17 and bacLP18) of two strains, Enterococcus mundtii LP17 and Enterococcus mundtii LP18, respectively, isolated from red mullet and sardine samples, determined large inhibition zones against all the Listeria species. Virulence traits and antibiotic resistances of all producers were verified, and no isolates presented dangerous characteristics, including the two best bacteriocin producers E. mundtii LP17 and E. mundtii LP18, which were subsequently investigated for their potential use in fish and seafood products biopreservation. For both strains, the highest level of bacteriocin production (1280 AU/ml) was recorded when cells were grown at 30 °C in MRS broth at pH ranging from 6.0 to 9.0, and high levels of adsorption of bacteriocins, bacLP17 and bacLP18, to the target cells Listeria monocytogenes were also observed. The results obtained in this study revealed that two strains of E. mundtii originating from seafood exhibited a strong inhibitory activity against L. monocytogenes and may be useful in controlling the growth of this pathogen in the same food products.

Volatile terpenes represent the largest group of Cannabis sativa L. components and they are responsible for its aromatic properties. Even if many studies on C. sativa have been focused on cannabinoids, which are terpenophenolics, little research has been carried out on its volatile terpenic compounds. In the light of all the above, the present work was aimed at the chemical characterization of seventeen essential oils from different fibre-type varieties of C. sativa (industrial hemp or hemp) by means of GC-MS and GC-FID techniques. In total, 71 compounds were identified, and the semi-quantitative analysis revealed that α- and β-pinene, β-myrcene and β-caryophyllene are the major components in all the essential oils analysed. In addition, a GC-MS method was developed here for the first time, and it was applied to quantify cannabinoids in the essential oils. The antibacterial activity of hemp essential oils against some pathogenic and spoilage microorganisms isolated from food and food processing environment was also determined. The inhibitory effects of the essential oils were evaluated by both the agar well diffusion assay and the minimum inhibitory concentration (MIC) evaluation. By using the agar diffusion method and considering the zone of inhibition, it was possible to preliminarily verify the inhibitory activity on most of the examined strains. The results showed a good antibacterial activity of six hemp essential oils against the Gram-positive bacteria, thus suggesting that hemp essential oil can inhibit or reduce bacterial proliferation and it can be a valid support to reduce microorganism contamination, especially in the food processing field.

Several species of the genus Acanthamoeba cause human diseases. Treatment of infections involves various problems, emphasising the need to develop alternative antiprotozoal agents. We studied the anti-amoebic activity of Essential Oils (EOs), derived from rosemary (Rosmarinus officinalis L.) and cloves (Syzygium aromaticum L. Merr. & Perry), against Acanthamoeba polyphaga strain. The amoebicidal activity of cloves and rosemary EOs was preliminary demonstrated by the morphology change (modifications in the cell shape, the presence of precipitates in the cytoplasm, autophagic vesicles, membrane blends) of the treated trophozoites. The cell-counts, carried out after staining trophozoites with a Trypan blue solution, revealed that both EOs were active in a dose-dependent manner and in relation to the exposure time. This activity was evident after few hours, with encouraging results obtained in particular with cloves EO, able to act at the lower concentrations and after 1 h, probably for its high eugenol content (65.30%).

Nisin, enterocin 416K1 and living bacteriocin-producer Enterococcus casseliflavus IM 416K1 have been entrapped in polyvinyl alcohol (PVOH) based coatings applied to poly (ethylene terephthalate) (PET) films, and their effectiveness in the control of the growth of Listeria monocytogenes ATCC 19117 has been tested. The anti-listerial activity of the doped coated films was evaluated by both a modified agar diffusion assay and a direct contact with artificially contaminated precooked chicken fillets stored at 4 °C, 22 °C and under simulated cold chain break conditions (1 day at 30 °C). The live-Enterococcus-doped film showed a more remarkable activity than nisin- and enterocin-doped films over long times both at 4 °C and 22 °C. The use of this film at 22 °C resulted in full inactivation of L. monocytogenes from the seventh day of the test. Live-Enterococcus-doped film displayed a much better antilisterial activity in comparison to nisin- and enterocin-doped films also in samples incubated at 4 °C, and submitted at one day (3rd or 7th day) of storage at 30 °C, to simulate cold chain break conditions. All results suggest that the live-Enterococcus-doped film can behave as a smart active food packaging, very effective in cold chain break conditions when the Listeria growth is fast., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

We investigated the occurrence of extended-spectrum β-lactamase (ESBL), AmpC, and carbapenemase-producing Gram-negative bacteria isolated from 160 samples of fresh vegetables (n = 80) and ready-to-eat (RTE) prepacked salads (n = 80). Phenotypic and genotypic analyses were carried out on the isolates in terms of the species present and relative resistance. Resistance to β-lactam antibiotics was found in only 44 (24 from fresh vegetables and 20 from RTE salads) of a total of 312 Gram-negative strains (14.1%). The prevalence of ESBL-producing strains from fresh vegetables was 83.3% (20/24) and 16.7% (4/24) for AmpC. Among the 20 bacterial isolates from RTE salads, 80% (16/20) were identified as ESBL-producing strains and the remaining 20% (4/20) as MBL-producing strains. PCR and sequencing confirmed the presence of bla SHV-12 , bla CTX-M-1 , bla CTX-M-15, bla RHAN -1 , bla ACC-1 , bla DHA-1 , bla VIM-1 , and bla IMP-1 . Seven different replicons were identified, where IncHI1, FIA, and I1 were the most representative types; when compared with the Inc types, isolates from fresh vegetables and RTE salads were similar. The location of genes on a conjugative plasmid was confirmed by positive results obtained with conjugation assays. Our study has demonstrated the occurrence and distribution of ESBL/AmpC and MBL strains in fresh vegetables and RTE salads in Italy and possible public health risks associated with consumption of these fresh products.

Purpose: Human leukocyte antigen (HLA)-G is a non-classic major histocompatibility complex HLA class I molecule. HLA-G may have tolerogenic properties which are linked to epigenetic-sensitive pathways. There is a correlation of sHLA-G levels and graft acceptance in transplantation studies. There are previous data on correlation of sHLA-G with graft rejection as well as with viral infections such as hepatitis C virus (HCV) in kidney transplanted patients. Here, we report the sHLA-G expression in patients on the waiting list for kidney transplantation, with and without anti-HCV compared to a control group., Methods: Serum of 67 patients on the waiting list for kidney transplantation (n = 43 with anti-HCV and n = 24 without anti-HCV) was analyzed. Among these patients, n = 39 were on the waiting list for the first transplantation, while n = 28 were patients who returned in the list. The control group included n = 23 blood donors with anti-HCV (n = 13) and without anti-HCV (n = 10)., Results: The expression of sHLA-G was significantly lower in the control group (39.6 ± 34.1 U/ml) compared to both - patients on the waiting list for the first transplantation (62.5 ± 42.4 U/ml, p=0.031) and patients who returned in the list (76.7 ± 53.9 U/ml, p=0.006). No significant differences were observed in all anti-HCV positive groups. A positive linear correlation between sHLA-G and TNF-α, and patient age was observed., Conclusions: Serum sHLA-G values were significantly increased in both - patients on the waiting list for the first transplantation and patients who returned in the list, as compared to control group. Our findings confirm the key tolerogenic role of sHLA-G levels as epigenetic-related marker for measuring the state of kidney allograft acceptance., (Copyright © 2018 Medical University of Bialystok. Published by Elsevier B.V. All rights reserved.)

Washing is a rapid and effective treatment to remediate contaminated sands impacted by oil spills, although synthetic additives used to increase extraction efficiency may cause additional pollution issues due to their intrinsic toxicity and very often low biodegradability. In this study, different biogenic mobilizing agents (soybean lecithins, cyclodextrins, cholic acids, plant-derived cleaners, rhamnolipids and sophorolipids) were tested in the washing of beach sands artificially contaminated with the Intermediate Fuel Oil IFO-180. Among these, a de-oiled soybean lecithin (SL-1), hydroxypropyl-β-cyclodextrins (HPB-CD) and sophorolipids (SR) achieved hydrocarbon removals close to those attained with the synthetic surfactant Triton™ X-100 (TX) in preliminary washing tests carried out at constant mixing rate, water/sand ratio and IFO-180 contamination level using agents concentrations close to their critical micelle concentration (0.1% and 1% w/v for microbial and non-microbial agents, respectively). The effects of agent concentration, water/sand ratio, mixing rate and IFO-180 contamination on hydrocarbons removal were modelled using face-centred central composite design and ANOVA. Optimal washing parameters for sand contamination levels in the range 0.5-20 g/kg were identified with response surface methodology. While HPB-CD and SR performed equally to TX only at low sand contaminations, SL-1 attained hydrocarbon removal higher or equal to that of TX at any IFO-180 contamination and at lower application rates. SL-1 also outperformed TX when minimizing the water/sand ratio, i.e., the volume of water used. Considering its lower toxicity, higher biodegradability and higher hydrocarbon removal efficiencies, SL-1 is an effective and environmentally sustainable alternative to synthetic surfactants in washing treatments for marine fuel-contaminated sands., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objectives: For decades, the detection of anti-HLA antibodies in candidates for solid-organ transplant has been performed with the traditional complement-dependent cytotoxicity method; this assay has been then integrated with the introduction of solid-phase assays. Over the past 20 years, the Luminex assay has become the most widely used in clinical laboratories due to both increased sensitivity and specificity versus enzyme-linked immunosorbent assay. However, even the Luminex technique has shown some critical issues, and choosing the most reliable method still remains challenging. In this study, we verified the concordance of the results obtained in detecting anti-HLA antibodies with 2 kit vendors that provide reagents for the Luminex platform., Materials and Methods: We used 314 serum samples from patients on wait lists for solid-organ transplant. Sera were tested with LABScreen Mixed-LSM12 (One Lambda-Thermo Fisher, Canoga Park, CA, USA) and LIFECODES LifeScreen Deluxe-LMX (Gen-Probe-Immucor, Stanford, CT, USA),which we indicated as vendor A and vendor B, respectively. Anti-HLA class I and class II antibody analyses were conducted by verifying the concordance of the results with Cohen kappa coefficient statistics and confidence interval., Results: The kappa coefficient statistics showed "substantial" reliability for class I (0.61; confidence interval, 0.50-0.73) and "moderate" reliability for class II (0.56; confidence interval, 0.43-0.69). There were no considerable differences in results between the 2 kits regarding overall assignment of negativity or positivity of a sample. Discordant data between positive values for a test and negative for the other were found for samples with weak antibody positivity., Conclusions: Some discordant data were probably attributable to several factors such as the composition of the kits, the antibody titer in the serum, whether sera were diluted, different washing methods, and type of plate used.

Heart transplantation is still the only possible life-saving treatment for end-stage heart failure, the critical epilogue of several cardiac diseases. Epigenetic mechanisms are being intensively investigated because they could contribute to establishing innovative diagnostic and predictive biomarkers, as well as ground-breaking therapies both for heart failure and heart transplantation rejection. DNA methylation and histone modifications can modulate the innate and adaptive immune response by acting on the expression of immune-related genes that, in turn, are crucial determinants of transplantation outcome. Epigenetic drugs acting on methylation and histone-modification pathways may modulate Treg activity by acting as immunosuppressive agents. Moreover, the identification of non-invasive and reliable epigenetic biomarkers for the prediction of allograft rejection and for monitoring immunosuppressive therapies represents an attractive perspective in the management of transplanted patients. MiRNAs seem to fit particularly well to this purpose because they are differently expressed in patients at high and low risk of rejection and are detectable in biological fluids besides biopsies. Although increasing evidence supports the involvement of epigenetic tags in heart failure and transplantation, further short and long-term clinical studies are needed to translate the possible available findings into clinical setting., (Copyright © 2017 Elsevier Inc. All rights reserved.)

HBV still represents a global risk factor in transfusion medicine. The residual risk of HBV is not limited to pre-seroconversion window period but it extends to donors with occult HBV infection (OBI) characterized by the presence of HBV DNA in liver and by the absence of the virus surface antigen. Each country developed an appropriate blood screening policy according to local HBV prevalence, yields of infectious units per different screening methods and cost-effectiveness. We underline the need of maintaining a high level of attention for OBI carrier identification in all blood banks worldwide where the screening procedures are generally based on a combination of both serological markers and nucleic acid amplification test. In this context, markers such as hepatitis B surface antibodies and hepatitis B core antibodies (anti-HBc) might be useful, although the use of this latter is highly debated and still controversial. Our aim is to give an overview on the relevant diagnostic approaches for the routine screening for HBV focusing on the feasibility of anti-HBc testing as precautionary measure in preventing OBI transmission worldwide. In our tailored algorithm, the loss of about 1% of 'anti-HBc only' donors, does not significantly affect the blood supply while improving recipient safety.

Serological assays are still considered the most useful tests in the diagnosis of syphilis. Since no single serological assay is able to provide a satisfactory result, in our laboratory we have evaluated the usefulness of a commercially-available immunoblot to diagnose syphilis infection among blood donors. From October 2012 to June 2013, 4572 blood donors were screened for syphilis with an automated chemiluminescent microparticle immunoassay (CMIA). To confirm the presence of treponemal antibodies, CMIA-reactive sera were tested by standard Treponema pallidum haemagglutination assay (TPHA). In addition, an alternative confirmatory test - the immunoblot INNO-LIA assay was introduced in our laboratory. Since two additional positives among CMIA-reactive-TPHA-negative samples were found, we concluded that the INNO-LIA immunoblot allowed a better detection of syphilis compared to TPHA. A confirmatory strategy based on the use of two treponemal assays could meet the screening requirements for blood donors as well as in our centre., (© The Author(s) 2015.)

Background: Alloimmunization remains a critical factor which affects the success of kidney transplantation. Patients awaiting solid organ transplantation may develop anti-HLA antibodies after pregnancies, transfusions and previous events of transplantations., Aim: We evaluated the effects of different sensitizing events on the anti-HLA antibody production and the potential role of patient HLA alleles in the context of antibody development in both the overall and pregnancy sensitized groups., Material and Methods: We retrospectively stratified 411 women on waiting list for kidney transplantation by route of sensitization. The presence of anti-HLA antibodies was evaluated by Solid Phase Assay and HLA typing was performed by serological and molecular methods., Results: In our study population, 54% of women had anti-HLA antibodies. We found that the 51.6% of women with pregnancy only, 44% of women with transfusion only and 100% of women with a history of transplantation only developed anti-HLA antibodies. Pregnancy only resulted significantly associated with all anti-HLA antibody development such as anti-A, -B, -C, -DR, -DP as well as anti-DQB and -DQA antibodies. We investigated the influence of patient HLA alleles on the antibody development in the overall study population. Patients expressing HLA A*32 (p=0.024, OR=0.42), B*14 (p=0.035, OR=0.44), HLA-B*44 (p=0.026, OR=0.51) and DRB1*01 (p=0.029, OR=0.55) alleles produced anti-HLA antibodies less frequently compared to subjects with other alleles. In the pregnancy only group, B*14 (p=0.010, OR=0.12) and B*51 (p=0.005, OR=0.24) alleles were associated with a low risk of anti-HLA antibody development, while A*11 (p=0.033, OR=3.56) and DRB1*04 (p=0.022, OR=3.03) alleles seem to represent a higher risk., Conclusions: Pregnancy still remains a strong sensitizing event in women awaiting kidney transplantation. The anti-HLA antibody development in pregnancy appears to be associated with the expression of particular HLA alleles., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The recrudescence of syphilis is leading to the development of new serological tests. The goal of this study was to compare the performance of the more recent Elecsys Syphilis assay, the Electro Chemiluminescence Immunoassay (ECLIA), with the former Architect Syphilis TP assay, the Chemiluminescent Microparticle Immunoassay (CMIA), for the detection of antibodies against Treponema pallidum in blood donors. Serum samples of 5543 voluntary blood donors were screened in parallel with two tests. All repeatedly reactive (RR) samples by one or both assays were further analysed for confirmation by immmunoblot INNO-LIA and TPHA. Of 32 RR samples by CMIA, 21 were confirmed positive; of 21 RR samples by ECLIA, 20 were confirmed positive. The sensitivities of CMIA and ECLIA were 100% and 95.24% (95% CI = 85.71-100), respectively, not significant (p > 0.05). The specificity and predictive positive value (PPV) of CMIA were 99.86% (95% CI = 99.74-99.94) and 72.41%, respectively, while the specificity and PPV of ECLIA were both 100%, being statistically significant (p = 0.01 for both). The overall agreement was 99.80% and the Cohen's kappa coefficients was 0.79. In conclusion, the recent Elecsys Syphilis assay could represent another reliable assay for blood donor screening.

Among several alternatives to control hospital-acquired infections (HAIs), a strategy could be the use of hospital uniforms imbued with antimicrobial substances. For this purpose we evaluated the antibacterial activity of two different silver doped fabrics employed for the production of hospital uniforms. The study was conducted in two-step. In the first the antimicrobial activity was evaluated in vitro against Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, Staphylococcus aureus ATCC 6538, Enterococcus faecalis ATCC 29212. In the second, we tested the total viable counts detected from beginning to end of the work shift on experimental silver doped uniforms worn by doctors, nurses, allied health assistants in different hospital wards. The in vitro tests showed a remarkable antibacterial activity of both silver doped samples (>99.9% reduction within 4h of exposure for Gram-positive and within 24 h for Gram-negative bacteria). The experimental uniforms provided results only slightly in agreement with in vitro data. Even if the increase of total viable counts was somewhat lower for experimental uniforms than traditional ones, significant differences were not observed. Despite the results on the uniforms worn, the addition of silver in fabrics to make medical equipment (supplies) remains an interesting option for HAI control.

We investigated presence and prevalence of antibiotic-resistances and other biological characters in enterococci isolated from faeces of healthy dogs and cats because these microorganisms represent important human and veterinary pathogens/opportunists, and a significant burden for healthcare systems. In all samples (n=115) we detected enterococci, with a predominance of Enterococcus faecium (42; 36.5%) and Enterococcus faecalis (36; 31.3%) species, endowed with virulence traits and multidrug-resistance. The two predominant resistance patterns (erythromycin, tetracycline) were examined by polymerase chain reaction for tet and erm genes. Only tetM for tetracycline, and ermA and ermB for erythromycin were detected. PCR for gelatinase gene (gelE) was positive in 62.6% of isolates, but only 26.1% produce gelatinase suggesting the existence of silent genes. efaAfs and efaAfm genes were found in E. faecalis and E. faecium respectively. 89.6% of isolates produced bacteriocin-like substances with a prevailing action against Listeria genus and, among these, 33.9% were positive for the bacteriocin structural genes entA, entL50 or entP. According to our study, pet animals can be considered a reservoir of potentially pathogenic enterococci and we cannot exclude that those microorganisms may be responsible for opportunistic infections in high-risk pet owners.

Recently, management of patients awaiting solid organ transplantation has taken advantages after the development of more sensitive and accurate solid phase assays which have supported the historic complement dependent cytotoxicity. This approach has allowed the detection of antibodies in patients previously considered negative. The use of the single antigen beads resulted in a more accurate anti-human leukocyte antigen (HLA) antibody characterization. The detection of anti-HLA antibodies specific for C, DQ and DP loci that were not so well characterized has been possible through the implementation of the single antigen assay. The assessment of HLA compatibility has been expanded through the introduction of "epitope matching" concept and the definition of the unacceptable antigens for a more adequate evaluation of donor-recipient compatibility. However, the clinical impact of pre-formed and de novo anti-HLA antibodies detected by solid phase assays is still controversial due to the drawback related to result interpretation. Until today, the unresolved issues concern if all antibodies affect the medium and long term clinical outcome. An open debate on the clinical relevance of anti-HLA antibodies detected by single-antigen beads highlights needing to further investigations. Here, we describe the novel applications and the improvements of the solid-phase assay use., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The aim of this study was to assess the production of extended spectrum beta-lactamases (ESBL) in 56 strains of Enterobacteriaceae, obtained from 100 rectal swabs of farm animals, and to evaluate the horizontal transfer ca- pacity of the genetic determinants of resistance. The ESBL-positive strains were confirmed by phenotypic testing, confirmed by PCR and DNA sequence analysis. The localization of beta-lactamase genes was established by conju- gation experiments. Of the 56 analyzed strains, 20 (36%) resulted positive for ESBL production by the double-disk synergy test, and belonged to Escherichia coli 15 (75%) and Klebsiella ozaenae 5 (25%) species. Molecular analysis showed that all ESBL-producing isolates possessed genes encoding for TEM-type enzymes and/or CTX-M. The conjugation assays yielded positive results, thus denoting a plasmidic localization of the genes. This study high- lights the high percentage of ESBL-positive Enterobacteriaceae and the mobility of the responsible genes. Gene mo- bility implies highly negative consequences in terms of drug therapy because of the spread of antibiotic resistance.

The aim of this study was to investigate the effects of different species of Lactobacilli on hyphal formation and biofilm development by the opportunistic fungal pathogen Candida albicans. We employed 4 different Lactobacillus species, namely L. rhamnosus, L. acidophilus, L. plantarum and L. reuteri, and 2 C. albicans strains, the reference DAY286 and its isogenic hwp1/hwp1 mutant, the FJS24 strain. As assessed by morphological analysis and quantitative colorimetric assays, Lactobacillus crude filtrate supernatant fluids (CFSF) affected Candida, impairing both hyphal formation and biofilm production. The CFSF-mediated phenomenon occurred in a dilution- and time-dependent manner and was consistently observed, irrespective of the C. albicans HWP1 genotype.

Automated chemiluminescent immunoassays (CLIAs) are useful for the detection of hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus 1/2 antigen/antibodies (HIV 1/2 Ag/Ab) in blood donor screening. Eight hundred and forty serum samples were tested for hepatitis B surface antigen (HBsAg), HCV antibodies (anti-HCV), and HIV1/2 Ag/Ab in parallel using 2 different CLIAs (Abbott Architect i2000SR and Roche Cobas e411). The concordance between the 2 systems was high (Cohen's kappa 0.97 for HBsAg, 0.77 for anti-HCV, 0.92 for HIV1/2 Ag/Ab) and the specificity and the positive predictive value were comparable. Among the 12 discrepant results, 11 were false-positive and 1 (reactive by Architect) was true-positive for anti-HCV. Positivity for HBV DNA, HCV RNA, and HIV RNA was recorded in 90.9%, 38.9%, and 100% of true-positive samples, respectively. This study represents the first stringent comparison between Architect i2000SR and Cobas e411 in blood donors. We observed a good correlation and high agreement among HBV, HCV, and HIV with the 2 automated systems.

Lactobacilli (150) from human vaginal secretions were tested for the production of antimicrobial substances which can provide a physiological defense against the pathogenic microorganisms in the vaginal area. Sixteen of the isolates (10.6%) showed antibacterial activity against one or several closely related microorganisms used as indicators. Lactobacillus fermentum CS57 was the best producer and secretes a bacteriocin-like substance (BLS) with antagonistic activity against Streptococcus agalactiae and Candida albicans. The compound was susceptible to the proteolytic enzymes and was heat labile. The mode of action was identified as bactericidal. The crude activity of the L. fermentum CS57 BLS was linked to a substance with a molecular weight larger than 30 kDa. Plasmid analysis of L. fermentum CS57 revealed the presence of a plasmid band with molecular weight of 54.7 kb. All L. fermentum CS57 non-producer variants (BLS-) obtained by curing experiments, showed loss of plasmid band and were susceptible to the BLS of the original strain. Therefore antimicrobial activity and immunity production seem to be linked to genes located on that same plasmid. Taking into account our results, L. fermentum CS57 could be considered a candidate for potential use as probiotic for the prophylaxis of vaginal human infections., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Recent evidences have shown that several host genetic factors influence susceptibility or protection to hepatitis C virus (HCV) infection. There are controversial data regarding the associations of human leukocyte antigens (HLA) and the clearance or progression of HCV. The aim of this study was to investigate whether particular HLA molecules were associated with HCV infection in recipients awaiting kidney transplantation considered at high-risk to infection due to protracted hemodialysis treatment. To this purpose, 301 kidney recipients with HCV infection and 1103 uninfected recipients were examined for HLA class I and II molecules. In our case-control study, HLA-A(*)26 is positively associated with HCV infection while HLA-A(*)29, -B(*)40 and -DRB1(*)01 are negatively associated with HCV infection. Multiple logistic regression analysis demonstrated that age (OR = 1.02; 95% CI = 1.01-1.04; p < 0.00), HLA-A(*)26, -A(*)29, -B(*)40 and -DRB1(*)01 [(OR = 1.54; 95% CI = 1.03-2.30; p = 0.03); (OR = 0.50; 95% CI = 0.26-0.99; p = 0.05); (OR = 0.42; 95% CI = 0.23-0, 7; p = 0.01); (OR = 0.62; 95% CI = 0.41-0, 94; p = 0.03); respectively] are independent predictors of HCV infection. Our results suggest that particular HLA molecules, as host genetic factors, may have a relationship with susceptibility or protection to HCV infection also in recipients awaiting kidney transplantation., (Copyright © 2013 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

In order to assess the frequency of clinically relevant linezolid-resistant staphylococcal isolates, and the role of linezolid in maintaining and coselecting multiple resistance mechanisms (cfr, 23S rRNA, L3/L4 mutations), a prospective Italian study was performed from 2010 to 2011 to confirm the diffusion of three major multidrug-resistant clones (ST2, ST5, ST23).

Considering the limited knowledge about the biological characters in enterococci isolated from surface waters, we investigated antibiotic and heavy-metal resistance, bacteriocin production, and some important virulence traits of 165 enterococci collected in water samples from Monte Cotugno Lake, the largest artificial basin built with earth in Europe. The species distribution of isolates was as follows: Enterococcus faecium (80%), Enterococcus faecalis (12.7%), Enterococcus casseliflavus (3%), Enterococcus mundtii (1.8%), Enterococcus hirae (1.8%), Enterococcus durans (0.6%). All enterococci showed heavy metal resistance toward Cu, Ni, Pb and Zn, were susceptible to Ag and Hg, and at the same time exhibited in large percentage (83.7%) resistance to one or more of the antibiotics tested. Relatively to virulence factor genes, 50.9% enterococci were positive for gelatinase (gelE), 10.9% for aggregation substance (agg), 12.7% and 66.6% for the cell wall adhesins (efaAfs and efaAfm), respectively. No amplicons were detected after PCR for cytolysin production (cylA, cylB and cylM) and enterococcal surface protein (esp) genes. Bacteriocin production was found in most of the isolates. Given that the waters of the Monte Cotugno Lake are used for different purposes, among which farming and recreational activities, they can contribute to spread enterococci endowed with virulence factors, and antibiotics and heavy metals resistance to humans.

The endosymbiotic relationship could represent for many bacteria an important condition favouring their spread in the environment and in foods. For this purpose we studied the behaviour of some food-borne and opportunistic pathogens (Listeria monocytogenes, Staphylococcus aureus, Enterococcus faecalis, Salmonella enterica serovar Enteritidis, Aeromonas hydrophila, Yersinia enterocolitica) when internalized in Acanthamoeba polyphaga. Our results confirm the capability of the bacteria tested to grow within amoebal hosts. We can observe two types of interactions of the bacteria internalized in A. polyphaga. The first type, showed by Y. enterocolitica and A. hydrophila, was characterized by an early replication, probably followed by the killing and digestion of the bacteria. The second type, showed by E. faecalis and S. aureus was characterized by the persistence and grow inside the host without lysis. Lastly, when amoebae were co-cultured with L. monocytogenes and S. Enteritidis, an eclipse phase followed by an active intracellular growth was observed, suggesting a third type of predator-prey trend. The extracellular count in presence of A. polyphaga, as a result of an intracellular multiplication and subsequent release, was characterized by an increase of E. faecalis, S. aureus, L. monocytogenes and S. Enteritidis, and by a low or absent cell count for Y. enterocolitica and A. hydrophila. Our study suggests that the investigated food-borne and opportunistic pathogens are, in most cases, able to interact with A. polyphaga, to intracellularly replicate and, lastly, to be potentially spread in the environment, underlining the possible role of this protozoan in food contamination., (Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

This work describes a simple and practical double synergy differential test (DSDT) that couples the detection of ESBLs and AmpC-type enzymes by means of a combo-disk approach using cefotaxime and ceftazidime as indicator substrates, and clavulanate and boronic acid as enzyme inhibitors. The DSDT was tested with a collection of 118 Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis strains with different beta-lactamase profiles, and proved to be highly sensitive and specific for the detection of ESBL and AmpC-producing isolates.

We describe here the isolation of 8 beta-lactamase-producing multidrug-resistant Enterococcus faecium isolates in 2010. All strains showed diverse pulsed-field gel electrophoresis (PFGE) profiles, all belonging to the same clonal complex, CC17. By PCR and hybridization experiments, the entire blaZ-blaI-blaR1 operon was found. The beta-lactamase activity was demonstrated at a high inoculum and in the presence of methicillin after overnight incubation.

Background: Staphylococcus aureus is a leading cause of community-acquired infections and healthcare-associated infections. Epidemiological data are useful for understanding the dynamics of the diffusion of this pathogen, and to plan control activities and monitor their efficacy., Methods: S. aureus isolates were collected in 13 public hospital laboratories of Emilia-Romagna (northern Italy region) during February-March 2009; phenotypic and molecular characterizations of these isolates were performed., Results: The study sample included 267 isolates, 57 from blood, 81 from respiratory tract, and 129 from wounds; 106 (40%) were methicillin-resistant S. aureus (MRSA). MRSA showed a limited number of circulating clones with 2 predominant spa types--t008 and t041--accounting for 36% and 27% of MRSA isolates, respectively. The t041 type had a higher prevalence of antimicrobial resistance compared to other spa types and accounted for most of the retrieved hetero-vancomycin-intermediate S. aureus (h-VISA), while t008 was more frequently detected in non-hospital isolates. A higher degree of genetic diversity was observed in methicillin-susceptible S. aureus (MSSA), with no predominant clones and low prevalence of antimicrobial resistance. The occurrence of community-acquired MRSA infection appears to be rare in Emilia-Romagna., Conclusions: In contrast to previous studies reporting Italian data, t008 was the most frequent spa type among MRSA isolates in Emilia-Romagna. The prevalence of antimicrobial resistance of different MRSA spa types could influence their ability to cause infections with hospital onset. The presence of only 2 major MRSA clones circulating in Emilia-Romagna increases the chances that a regional strategy aimed at MRSA prevention will be effective.

In last decade methicillin-resistant Staphylococcus aureus with high level of vancomycin-resistance (VRSA) have been reported and generally the patients with VRSA infection were also infected with a vancomycin-resistant Enterococcus (VRE). Considering that the high level of vancomycin-resistance in VRSA isolates seems to involve the horizontal transfer of Tn1546 transposon containing vanA gene from coinfecting VRE strains, the authors have studied the "in vitro" conjugative transfer of this resistance from VanA enterococci to S. aureus. Out of 25 matings performed combining five vancomycin-resistant enterococci as donors (three Enterococcus faecalis and two Enterococcus faecium), and five S. aureus as recipients, all clinical isolates, two have been successful using E. faecalis as donor. The transfer of vancomycin-resistance was confirmed by vanA gene amplification in both transconjugants and the resistance was expressed at lower levels (MIC 32 μg/ml) in comparison with the respective VRE donors (MIC > 128 μg/ml). The vancomycin-resistance of trasconjugants was maintained even after subsequent overnight passages on MSA plates containing subinhibitory levels of vancomycin. This study shows that the vanA gene transfer can be achieved through techniques "in vitro" without the use of laboratory animals employed, in the only similar experiment previously carried out by other authors, as substrate for the trasconjugant growth. Moreover, in that previous experiment, contrary to this study, the vancomycin resistant S. aureus trasconjugants were selected on erythromycin agar and not by direct vancomycin agar selection.

This study aimed to evaluate the accuracy of routine systems (Vitek2 cards AST-N022 and AST-N026; Kirby Bauer; Etest) for susceptibility testing of Pseudomonas aeruginosa to piperacillin and piperacillin-tazobactam. Vitek2 (card AST-N022) showed the worst performance; the other three methods (Vitek2 card AST-N026, Kirby-Bauer and E-test) performed comparably but never fulfilled the minimal standard proposed by FDA.

Probiotic compounds, which are often constituted of lactobacilli, exert a number of health benefits through maintenance of the intestinal ecosystem balance. Among the important interactions that occur in the gut microbiota, plasmid transfer by mating is an increasing cause of concern, particularly when antibiotic-resistant genes are involved. Because lactobacilli seem to be able to influence this mechanism, the aim of the present work was to investigate the in vitro capability of two Lactobacillus plantarum strains (one bacteriocin producer and one nonproducer) to interfere with the conjugation processes. For this purpose different matings were performed adding to the donor and recipient cells L. plantarum 35d bac+ and L. plantarum 396/1 bac- as agents of interference. Conjugations added with a Staphylococcus aureus strain or without any agent of interference were used as controls. The results of our experiments demonstrated that both lactobacillus strains were able to decrease mating frequency. Statistically significant differences in the viable transconjugants were obtained in the presence and in the absence of the lactobacilli. The effect was almost the same with the two L. plantarum independent of bacteriocin production. In the trial performed with S. aureus, no decrease in mating frequency was observed, confirming that the capability to interfere with R-plasmid transfer ability could be a property of the tested L. plantarum strains.