Antiviral Activity of Reagents in Mouth Rinses against SARS-CoV-2.

The oral cavity, an essential part of the upper aerodigestive tract, is believed to play an important role in the pathogenicity and transmission of SARS-CoV-2. The identification of targeted antiviral mouth rinses to reduce salivary viral load would contribute to reducing the COVID-19 pandemic. While awaiting the results of significant clinical studies, which to date do not exist, the commercial availability of mouth rinses leads us to search among them for reagents that would have specific antiviral properties with respect to SARS-CoV-2. The challenges facing this target were examined for 7 reagents found in commercially available mouth rinses and listed on the ClinicalTrials.gov website: povidone-iodine, chlorhexidine, hydrogen peroxide, cyclodextrin, Citrox, cetylpyridinium chloride, and essential oils. Because SARS-CoV-2 is an enveloped virus, many reagents target the outer lipid membrane. Moreover, some of them can act on the capsid by denaturing proteins. Until now, there has been no scientific evidence to recommend mouth rinses with an anti-SARS-CoV-2 effect to control the viral load in the oral cavity. This critical review indicates that current knowledge of these reagents would likely improve trends in salivary viral load status. This finding is a strong sign to encourage clinical research for which quality protocols are already available in the literature.

Biocatalytic Self-Assembly Using Reversible and Irreversible Enzyme Immobilization.

Biocatalytic control of molecular self-assembly provides an effective approach for developing smart biomaterials, allowing versatile enzyme-mediated tuning of material structure and properties as well as enabling biomedical applications. We functionalized surfaces with bioinspired polydopamine and polyphenol coatings to study the effects of enzyme surface localization and surface release on the self-assembly process. We show how these coatings could be conveniently used to release enzymes for bulk gelation as well as to irreversibly immobilize enzymes for localizing the self-assembly to the surface. The results provide insights to the mode of action of biocatalytic self-assembly relevant to nanofabrication and enzyme-responsive materials.

Metastable hydrogels from aromatic dipeptides.

We demonstrate that the well-known self-assembling dipeptide diphenylalanine (FF) and its amidated derivative (FF-NH2) can form metastable hydrogels upon sonication of the dipeptide solutions. The hydrogels show instantaneous syneresis upon mechanical contact resulting in rapid expulsion of water and collapse into a semi-solid gel.

Plasmid-mediated fluoroquinolone resistance determinants in Escherichia coli from community uncomplicated urinary tract infection in an area of high prevalence of quinolone resistance.

In Italy fluoroquinolones (FQs) are extensively prescribed in empirical therapy of uncomplicated urinary tract infection (UTI) despite recommendations in national guidelines and widespread antibiotic resistance in community. To survey the dissemination of plasmid-mediated quinolone resistance in a peak area of FQs consumption, E. coli strains from 154 community and 41 local hospital patients were collected; low level ciprofloxacin resistance qnrA, qnrB, qnrS, and aac(6)'-Ib-cr genes were screened by PCR and patterns of transferable resistances were determined. Clinical ciprofloxacin resistance in hospital doubled community value, while overall rates of FQ resistance genes were similar (31.6% and 27.8%). Prevalence of aac(6')-Ib-cr gene was 11% in outpatients (21%, inpatients) and risk of harbouring this variant was significantly associated with gentamicin resistance; linkage to ceftazidime resistance was significant (P=0.001) and six out of eight strains produced CTX-M-15 and TEM-1 beta lactamases. In transconjugants, the unique pattern ampicillin/kanamycin-gentamicin/ ESBL + was associated with aac(6')-Ib-cr gene presence and with an increase of ciprofloxacin MIC value. Data highlight the need to monitor the resistance risk factors in the local community to provide clinicians with well-grounded guidelines for UTI therapy.

JC Viral reactivation in a pediatric patient with Crohn's disease.

This is a report concerning human polyomavirus JC (JCV) reactivation in a pediatric patient with Crohn's disease (CD) during the treatment with 5-aminosalicylic acid (5-ASA), a non-steroidal anti-inflammatory drug (NSAID). We examined 9 bioptic samples from three different bowel districts (ileum, cecum, rectum) of this child. These samples were analyzed by Quantitative PCR (Q-PCR) to investigate the presence of JCV DNA. JCV DNA was detected in one rectum biopsy taken two months after 5-ASA treatment. Although our result must be validated in a larger group of subjects and with a longer follow-up period, it underlines the importance of JVC monitoring in CD patients.

Bacterial ratchet motors.

Self-propelling bacteria are a nanotechnology dream. These unicellular organisms are not just capable of living and reproducing, but they can swim very efficiently, sense the environment, and look for food, all packaged in a body measuring a few microns. Before such perfect machines can be artificially assembled, researchers are beginning to explore new ways to harness bacteria as propelling units for microdevices. Proposed strategies require the careful task of aligning and binding bacterial cells on synthetic surfaces in order to have them work cooperatively. Here we show that asymmetric environments can produce a spontaneous and unidirectional rotation of nanofabricated objects immersed in an active bacterial bath. The propulsion mechanism is provided by the self-assembly of motile Escherichia coli cells along the rotor boundaries. Our results highlight the technological implications of active matter's ability to overcome the restrictions imposed by the second law of thermodynamics on equilibrium passive fluids.

Virulence traits in Escherichia coli strains isolated from outpatients with urinary tract infections.

This study aims to characterize phenotypic and genotypic virulence traits in Escherichia coli strains, isolated from outpatients with urinary tract infections, comparing with those obtained from inpatients. Information on the pathogenic behavior of the uropathogenic strains was obtained by monitoring different biological properties, such as autoagglutination, hemagglutination, adhesiveness to and invasion of human bladder (HT1376) cells, biofilm formation, phylogenetic grouping, and virulence-related genes. The results show similar behavior in the two groups concerning autoagglutination, hemagglutination, and biofilm formation. None of the strains examined was invasive. However, in strains from outpatients there was an increased adhesion to HT1376 cells compared with clinical strains, a significant higher presence of genes codifying for adhesins and cell protection factors, and a lower proportion of strains belonging to B1 group. These findings add further information on the pathogenic traits of community E. coli, since strains isolated from the outpatients' group were differently "armed" in comparison with those of clinical cases, and more suitable to infect healthy individuals.

Invasive pathway of Listeria ivanovii in human amnion-derived WISH cells.

Among Listeria genus, only two species, Listeria ivanovii and Listeria monocytogenes, are pathogenic. L. ivanovii is almost only associated with infections in animals, mainly sheep and cattle, and has rarely been associated with human infections, whereas L. monocytogenes causes severe illnesses in both humans and animals. To further investigate the pathogenetic features of L. ivanovii in humans, we undertook a study in which the intracellular behaviour of this pathogen was analysed in WISH cells, a cell line derived from human amniotic tissue, and compared to that of L. monocytogenes. Using microbiological, biochemical, and ultrastructural approaches, we demonstrate that L. ivanovii can adhere to and invade human amniotic cells, lyse the phagosomal membrane, polymerize host cell actin, and spread from cell to cell more efficiently than L. monocytogenes. However, although L. ivanovii is capable of specifically infecting and replicating in human amnion cells, its survival in cytoplasm is limited compared to that of L. monocytogenes.

Acid adaptation and survival of Listeria monocytogenes in Italian-style soft cheeses.

AIMS: The ability of Listeria monocytogenes to survive and grow at high salt concentrations and low pH makes it a potential hazard after the consumption of milk and dairy products, often implicated in severe outbreaks of listeriosis. This study was designed to evaluate the behaviour of L. monocytogenes in traditional acid and salted Italian-style soft cheeses and to investigate whether Listeria occurrence and growth in these environments may represent a potential increase of hazard. METHODS AND RESULTS: A first approach was addressed to in vitro evaluate survival, acid tolerance response, ability to produce biofilm, and capability to invade intestinal-like cells of a L. monocytogenes strain grown under experimental conditions mimicking environmental features that this pathogen encounters in soft cheeses (such as acid pH and high NaCl content). A second set of experiments was performed to monitor, during the storage at 4 degrees C, the survival of acid-adapted and nonadapted Listeriae in artificially contaminated soft cheeses. Both acid tolerance response and invasion efficiency of acid-adapted bacteria resulted in an increase, even when bacteria were simultaneously pre-exposed to increasing salt stress. The contamination of cheeses with acid-adapted and nonadapted bacteria evidenced in all products a good survival. A significant increased survival, the recovery of bacterial cells highly resistant to lethal pH exposure, and the prevalence of filamentous structures were observed in crescenza cheese during the storage. CONCLUSIONS: The Listeria survival and acid pH tolerance observed during refrigerated storage are probably related to the intrinsic acid and saline features of soft cheeses analysed. SIGNIFICANCE AND IMPACT OF THE STUDY: Italian soft cheeses tested may represent a potential hazard for the recovery of acid-adapted L. monocytogenes cells with enhanced ability to adhere to inert surfaces and/or to penetrate host cells.

Listeria monocytogenes in a young patient with non Hodgkins lymphoma: case report.

Listeria monocytogenes is an intracellular food-borne pathogen, widely distributed in the environment, which rarely causes clinical infection in healthy people, but may cause severe disease in immunocompromised patients. A case of listeriosis is certified in an immunocompromised patient, thus confirming this microorganism to be an opportunistic human pathogen.

Gut-associated bacterial microbiota in paediatric patients with inflammatory bowel disease.

BACKGROUND: Clinical and experimental observations in animal models indicate that intestinal commensal bacteria are involved in the initiation and amplification of inflammatory bowel disease (IBD). No paediatric reports are available on intestinal endogenous microflora in IBD. AIMS: To investigate and characterise the predominant composition of the mucosa-associated intestinal microflora in colonoscopic biopsy specimens of paediatric patients with newly diagnosed IBD. METHODS: Mucosa-associated bacteria were quantified and isolated from biopsy specimens of the ileum, caecum and rectum obtained at colonoscopy in 12 patients with Crohn's disease, 7 with ulcerative colitis, 6 with indeterminate colitis, 10 with lymphonodular hyperplasia of the distal ileum and in 7 controls. Isolation and characterisation were carried out by conventional culture techniques for aerobic and facultative-anaerobic microorganisms, and molecular analysis (16S rRNA-based amplification and real-time polymerase chain reaction assays) for the detection of anaerobic bacterial groups or species. RESULTS: A higher number of mucosa-associated aerobic and facultative-anaerobic bacteria were found in biopsy specimens of children with IBD than in controls. An overall decrease in some bacterial species or groups belonging to the normal anaerobic intestinal flora was suggested by molecular approaches; in particular, occurrence of Bacteroides vulgatus was low in Crohn's disease, ulcerative colitis and indeterminate colitis specimens. CONCLUSION: This is the first paediatric report investigating the intestinal mucosa-associated microflora in patients of the IBD spectrum. These results, although limited by the sample size, allow a better understanding of changes in mucosa-associated bacterial flora in these patients, showing either a predominance of some potentially harmful bacterial groups or a decrease in beneficial bacterial species. These data underline the central role of mucosa-adherent bacteria in IBD.

Encrusted cystitis in an immunocompromised patient: possible coinfection by Corynebacterium urealyticum and E. coli.

Encrusted cystitis is a severe chronic inflammatory disease of the bladder characterized by excessively alkaline urine and calcifications within the bladder wall. A case of a 60 year-old man affected by systemic lupus erythematosus (SLE), which developed encrusted cystitis due to Corynebacterium urealyticum with E. coli co-infection, shows that the treatment of encrusted cystitis with a endoscopic debulking of the encrusted stones and an antimicrobial therapy specific for C. urealyticum often is not sufficient for the complete resolution of symptoms.

Apoptotic death of Listeria monocytogenes-infected human macrophages induced by lactoferricin B, a bovine lactoferrin-derived peptide.

Listeria monocytogenes, an intracellular facultative food-borne pathogen, was reported to induce apoptosis in vitro and in vivo in a variety of cell types with the exception of murine macrophages. These cells represent the predominant compartment of bacterial multiplication and die as a result of necrosis. In this study we showed that human non-activated and IFN-gamma-activated macrophagic-like (THP-1) cells infected with L. monocytogenes, mainly die by necrosis rather than by an apoptotic process. Two natural products derived from bovine milk, lactoferrin and its derivative peptide lactoferricin B, are capable of regulating the fate of infected human macrophages. Bovine lactoferrin treatment of macrophages protects them from L. monocytogenes-induced death whereas lactoferricin B, its derivative peptide, determines a shifting of the equilibrium from necrosis to apoptosis.

Extended Spectrum Beta-Lactamase-producing Klebsiella pneumoniae outbreaks during a third generation cephalosporin restriction policy.

In spite of the adoption of third generation cephalosporin restriction policies, two independent outbreaks by Extended Spectrum Beta-Lactamase (ESBL)-producing Klebsiella pneumoniae occurred in two different wards (Neonatal Intensive Care Unit, NICU and Neurosurgery) of a teaching hospital in Rome, Italy. In the former 19 infected neonates were reported, whereas in the latter there were 10 infected patients. In both wards no differences were observed in the mortality rates in periods of outbreak and those with no outbreak. Molecular typing on a total of 19 isolated strains was carried out and restriction patterns were compared. The PFGE showed that nine isolates responsible for infection in the NICU were all included in three closely related clusters. In Neurosurgery nine strains out of ten were strictly related and part of an outbreak occurring between August-December 2003, while one isolate was temporarily (February 2003) and genetically (seven band differences) unrelated to the outbreak strains. When ESBL producing K. pneumoniae clusters from the two wards (NICU vs Neurosurgery) were compared, they appeared to be completely different both for their genotype pattern and plasmid type or presence, thus demonstrating cross transmission by two different genotypes.

Infectious agents in tissues from spontaneous abortions in the first trimester of pregnancy.

Some evidence suggests that intrauterine infection plays a major role in the pathogenesis of early pregnancy loss, but the implication and prevalence of microrganisms in the aetiology of spontaneous abortion during the first trimester of pregnancy has not yet been well established. In this study, we analysed the tissues relative to the product of conception from abortions during the first trimester (51 spontaneous abortions and 56 voluntary pregnancy interruptions) in women attending the Gynecological Sciences Perinatology and Puericulture Department of "Policlinico Umberto I". Specimens were investigated by cultural methods for the presence of yeasts, gram positive, gram negative bacteria, and genital mycoplasma. By molecular diagnostic procedures, DNA sequences of Chlamydia trachomatis, herpes simplex viruses, adenovirus, human papillomaviruses and human polyomaviruses BK and JC were searched. None of these agents could be found in voluntary pregnancy interruption samples, with the exception of 3.6% of specimens positive for adenovirus, whereas spontaneous abortion tissues were positive for at least one microrganism by 31.5%. Data analysis showed the occurrence of both monomicrobial and polymicrobial infections.

Heterogeneity of virulence-related properties in Listeria monocytogenes strains isolated from patients with haematological malignancies.

Listeria monocytogenes is an intracellular foodborne pathogen of humans and animals for which there are indications of virulence differences among strains. Various virulence properties related to different phases of infection process were investigated in L. monocytogenes strains isolated from patients affected by haematological malignancies. In these isolates, besides to the clinical history, we analysed the haemolysin production, the survival to acidic pH, the ability to enter and proliferate in human intestinal-like and human macrophagic-like cells, as well as the allelic polymorphism of the actA gene involved intracellular movement. A general heterogeneity in the virulence properties was detected which did not appear correlated with the clinical outcome of listeriosis but more probably was influenced by the status of the immune defence of the host.

Detection of Listeria monocytogenes in Italian-style soft cheeses.

AIMS: A rapid detection system specific for Listeria monocytogenes and based on the polymerase chain reaction (PCR) was developed. METHODS AND RESULTS: Primers annealing to the coding region of the actA gene, critically involved in virulence and capable of discrimination between two different alleles naturally occurring in L. monocytogenes, have been utilized. The procedure was applied to recover L. monocytogenes cells in artificially contaminated fresh Italian soft cheeses (mozzarella, crescenza and ricotta). Low levels of L. monocytogenes were detected in mozzarella and crescenza homogenates (0.04-0.4 and 4 CFU g(-1), respectively) whereas in ricotta the detection limit was higher (40 CFU g(-1)). CONCLUSIONS: This PCR-based assay is highly specific as primers used recognize the DNA from different L. monocytogenes strains of clinical and food origin, while no amplification products result with any other Listeria spp. strains. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlighted a low-cost and rapid procedure that can be appropriated for the detection in real time of low L. monocytogenes levels in soft cheese.

Involvement of herpes simplex type 2 in modulation of gene expression of human papillomavirus type 18.

Human papillomaviruses (HPVs) and herpes simplex virus type 2 (HSV-2) can establish latent or persistent infections in the host, and are involved in the aetiology of benign and/or malignant lesions of the urogenital tract. To investigate the putative interaction between these DNA viruses when a double infection occurs, we have studied the effect of HSV-2 infection in HeLa 229 cells containing 10-50 copies of HPV type 18 genomic DNA. Twenty hours post HSV-2 infection, the analysis of mRNA transcripts from E1, E2, E6 early and L1 late HPV18 genes was performed in HeLa cells by a semi-quantitative RT-PCR assay. A modulation of HPV18 E1 and E6 early genes was observed, resulting in a 9-fold and 3-fold increased transcription respectively.

Infection of human enterocyte-like cells with rotavirus enhances invasiveness of Yersinia enterocolitica and Y. pseudotuberculosis.

Mixed infection with rotavirus and either Yersinia enterocolitica or Y. pseudotuberculosis was analysed in Caco-2 cells, an enterocyte-like cell line highly susceptible to these pathogens. Results showed an increase of bacterial adhesion and internalisation in rotavirus-infected cells. Increased internalisation was also seen with Escherichia coli strain HB101 (pRI203), harbouring the inv gene from Y. pseudotuberculosis, which is involved in the invasion process of host cells. In contrast, the superinfection with bacteria of Caco-2 cells pre-infected with rotavirus resulted in decreased viral antigen synthesis. Transmission electron microscopy confirmed the dual infection of enterocytes. These data suggest that rotavirus infection enhances the early interaction between host cell surfaces and enteroinvasive Yersinia spp.

Acid tolerance in Listeria monocytogenes influences invasiveness of enterocyte-like cells and macrophage-like cells.

Clinical and food Listeria monocytogenes isolates, pre-exposed to mild acidic conditions, were able to readily develop acid tolerance, irrespective of their origin. We attempted to investigate the influence of acid tolerance mechanisms, either constitutive or induced, on the invasive behaviour of this facultative food-borne pathogen. Entry efficiency and intracellular growth of acid-tolerant strains were evaluated in in vitro cell models capable to mimic in vivo target cells, such as enterocytes and macrophages. An acid-adapted L. monocytogenes wild-type strain and a constitutively acid-tolerant mutant were able to enter enterocyte-like (Caco-2) cells as well as to survive and proliferate intracellularly in lipopolysaccharide-treated macrophage-like (J774.A1) cells, at a significant increased extent by respect of the non acid-adapted wild-type strain. These findings add new information about the influence of the acid tolerance response on L. monocytogenes virulence, suggesting that in acid-adapted bacteria the early events of pathogenesis which allow the colonization and the spread of bacteria in the host may be highly promoted.