Immacolata Anacarso

Anti-listerial activity of a polymeric film coated with hybrid coatings doped with Enterocin 416K1 for use as bioactive food packaging.

In this study, Enterocin 416K1, a bacteriocin produced by Enterococcus casseliflavus IM 416K1, was entrapped in an organic-inorganic hybrid coating applied to a LDPE (low-density polyethylene) film for its potential use in the active food packaging field. The antibacterial activity of the coated film was evaluated against Listeria monocytogenes NCTC 10888 by qualitative modified agar diffusion assay, quantitative determination in listeria saline solution suspension and direct contact with artificially contaminated food samples (frankfurters and fresh cheeses) stored at room and refrigeration temperatures. All investigations demonstrated that enterocin-activated coatings have a good anti-listeria activity. Qualitative tests showed a clear zone of inhibition in the indicator lawn in contact with and around the coated film. During the quantitative antibacterial evaluation the L. monocytogenes viable counts decreased to 1.5 log units compared to the control. The inhibitory capability was confirmed also in food-contact assays. In all food samples packed with coated films we observed a significant decrease in L. monocytogenes viable counts in the first 24 h compared to the control. This difference was generally maintained up to the seventh day and then decreased, with the exception of the cheese samples stored at refrigeration temperature.

Detection of bacteriocin production and virulence traits in vancomycin-resistant enterococci of different sources.

Aim:  Three hundred and two enterococci were isolated from food, animal and clinical samples in order to evaluate the incidence of vancomycin‐resistant enterococci (VRE) and bacteriocin, cytolysin, haemolysin, gelatinase production.

Acanthamoeba polyphaga, a potential environmental vector for the transmission of food-borne and opportunistic pathogens

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. (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Ecological behaviour of three serogroups of Legionella pneumophila within a model plumbing system

Three Legionella pneumophila strains isolated from water samples and belonging to serogroups (sgs) 1, 6 and 9 were analysed for their capacity to colonise an experimental model simulating a domestic hot water distribution system. Ecological factors that could influence the persistence of the sgs such as intracellular life within protozoan hosts and bacterial interference by the production of antagonistic compounds were also studied. Viable counts of L. pneumophila increased both in the planktonic and in the sessile phases. Sg 6 showed a marked prevalence during the whole experiment and exhibited the highest host infection efficiency. Sg 1 was significantly less represented, but showed the highest capacity to reproduce in the protozoan hosts. Sg 9 was poorly represented and less adapted to intracellular life. Among the 14 bacteria constantly isolated in the system, five (35.7%) produced antagonistic substances against Legionella, with differences according to the bacterial strain and L. pneumophila sgs.

Protozoa and human macrophages infection by Legionella pneumophila environmental strains belonging to different serogroups.

Three Legionella pneumophila strains isolated from municipal hot tap water during a multicentric Italian survey and belonging to serogroups 1, 6, 9 and the reference strain Philadelphia-1 were studied to determine the intracellular replication capability and the cytopathogenicity in human monocyte cell line U937 and in an Acanthamoeba polyphaga strain. Our results show that both serogroups 1 and Philadelphia-1 were able to multiply into macrophages inducing cytopathogenicity, while serogroup 6 and ever more serogroup 9 were less efficient in leading to death of the infected macrophages. Both serogroups 1 and 6 displayed a quite good capability of intracellular replication in A. polyphaga, although serogroup 1 was less cytopathogenic than serogroup 6. Serogroup 9, like Philadelphia-1 strain, showed a reduced efficiency of infection and replication and a low cytopathogenicity towards the protozoan. Our study suggests that bacterial pathogenesis is linked to the difference in the virulence expression of L. pneumophila serogroups in both hosts, as demonstrated by the fact that only L. pneumophila serogroup 1 shows the contextual expression of the two virulence traits. Serogroup 6 proves to be a good candidate as pathogen since it shows a good capacity for intracellular replication in protozoan.Three Legionella pneumophila strains isolated from municipal hot tap water during a multicentric Italian survey and belonging to serogroups 1, 6, 9 and the reference strain Philadelphia-1 were studied to determine the intracellular replication capability and the cytopathogenicity in human monocyte cell line U937 and in an Acanthamoeba polyphaga strain. Our results show that both serogroups 1 and Philadelphia-1 were able to multiply into macrophages inducing cytopathogenicity, while serogroup 6 and ever more serogroup 9 were less efficient in leading to death of the infected macrophages. Both serogroups 1 and 6 displayed a quite good capability of intracellular replication in A. polyphaga, although serogroup 1 was less cytopathogenic than serogroup 6. Serogroup 9, like Philadelphia-1 strain, showed a reduced efficiency of infection and replication and a low cytopathogenicity towards the protozoan. Our study suggests that bacterial pathogenesis is linked to the difference in the virulence expression of L. pneumophila serogroups in both hosts, as demonstrated by the fact that only L. pneumophila serogroup 1 shows the contextual expression of the two virulence traits. Serogroup 6 proves to be a good candidate as pathogen since it shows a good capacity for intracellular replication in protozoan.

Anti-listerial activity of coatings entrapping living bacteria

Polyvinyl alcohol (PVOH) based coatings entrapping either living bacteriocin-producer Enterococcus casseliflavusIM 416K1 bacteria or Enterocin 416K1 have been prepared and applied to poly(ethylene terephthalate) (PET) films. The antimicrobial activity of coated PET films was evaluated against Listeria monocytogenes NCTC 10888 by qualitative agar diffusion assays and by direct contact with artificially contaminated food samples (wurstel and seasoned cheese) stored at 4 °C and 22 °C. Anti-listerial activity of both coatings was observed for both tests. However, the live-enterococcus doped coatings showed a much more remarkable anti-listerial activity than enterocin doped ones. Interestingly, live-enterococcus doped coatings lead to a strong decrease of L. monocytogenes viable counts even at 22 °C, indicating that they are able to contrast efficiently the fast L. monocytogenesgrowth occurring at this temperature in wurstel samples. In this respect, they can be considered smart coatings, being able to be responsive towards an accidental rise of temperature during food storage. The capability of bacteria to survive for a long time can also assure a long lasting antibacterial activity.

Antibiotics and heavy metals resistance and other biological characters in enterococci isolated from surface water of Monte Cotugno Lake (Italy)

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.

Conjugation-Mediated Transfer of Antibiotic-Resistance Plasmids Between Enterobacteriaceae in the Digestive Tract of Blaberus craniifer (Blattodea: Blaberidae)

Abstract Cockroaches, insects of the order Blattodea, seem to play a crucial role in the possible conjugation-mediated genetic exchanges that occur among bacteria that harbor in the cockroach intestinal tract. The gut of these insects can be thought of as an effective in vivo model for the natural transfer of antimicrobial resistance plasmids among bacteria. In our study, we evaluated the conjugation-mediated horizontal transfer of resistance genes between Escherichia coli and other microorganisms of the same Enterobacteriaceae family within the intestinal tract of Blaberus craniifer Burmeister, 1838 (Blattodea: Blaberidae). Different in vivo mating experiments were performed using E. coli RP4 harboring the RP4 plasmid carrying ampicillin, kanamycin, and tetracycline resistance genes as the donor and E. coli K12 resistant to nalidixic acid or Salmonella enterica serovar Enteritidis IMM39 resistant to streptomycin as the recipients. The RP4 plasmid was successfully transferred to both recipients, producing E. coli K12-RP4 and S. Enteritidis IMM39-RP4 transconjugants. Conjugation frequencies in vivo were similar to those previously observed in vitro. The transfer of the RP4 plasmid in all transconjugants was confirmed by small-scale plasmid isolation and agar gel electrophoresis, suggesting that the intestinal tract of cockroaches is an effective in vivo model for natural gene transfer. Our results confirm that cockroaches allow for the exchange of antimicrobial resistance plasmids among bacteria and may represent a potential reservoir for the dissemination of antibiotic-resistant bacteria in different environments. These findings are particularly significant to human health in the context of health care settings such as hospitals.

Amoebicidal Effects of Three Bacteriocin like Substances from Lactic AcidBacteria against Acanthamoeba Polyphaga

We investigated the antiamoebic activity of three Bacteriocin Like Substances (BLS 39, BLS GS 54, BLS GS 16) produced by Lactic Acid Bacteria (LAB). The crude bacteriocins showed an amoebicidal effect against Acanthamoeba polyphaga, but with differences. BLS 39, produced by Lactobacillus pentosus, determined a prompt and progressive decrease of viable amoebal cell count, up to the end of the experiment (144 h), where the trophozoites were not detectable. A killing effect, but after a more prolonged contact time, was observed for BLS GS 54, produced by Lactobacillus paraplantarum, whereas the bacteriocin produced by Lactobacillus plantarum GS16 showed the lowest toxicity for A. polyphaga. For BLS GS 16 the maximum percentage of reduction in trophozoites count (45%) was obtained after 144 h, value much lower when compared to BLS GS54 and BLS 39, that showed values of 44,60% and 52,60% after only one hour of contact, with a maximum of 98% and 100% of non-viable cells, respectively, after 144 h. Morphological changes of the A. polyphaga cells as swollen cells, roundness and cellular lysis, were already observed after the first hours of contact with BLS and, at the end of the experiment, most of the cells were colored (blue), indicating their death. Currently there isn

Culture Compounds which are Able to Increase the Growth and the Production of Bacteriocins by Two Different LABS

prime;t evidence of BLS produced by LAB active against amoebas. In this study we have shown that all the three BLS secreted by the Lactic Acid Bacteria are endowed with amoebicidal effect against Acanthamoeba polyphaga, killing the protozoan with different effectiveness and at different times of contact.