Risk factors for recurrence in patients with Clostridium difficile infection due to 027 and non-027 ribotypes.

OBJECTIVES: Our objective was to evaluate factors associated with recurrence in patients with 027+ and 027- Clostridium difficile infection (CDI). METHODS: Patients with CDI observed between January and December 2014 in six hospitals were consecutively included in the study. The 027 ribotype was deduced by the presence of tcdB, tcdB, cdt genes and the deletion Δ117 in tcdC (Xpert® C. difficile/Epi). Recurrence was defined as a positive laboratory test result for C. difficile more than 14 days but within 8 weeks after the initial diagnosis date with reappearance of symptoms. To identify factors associated with recurrence in 027+ and 027- CDI, a multivariate analysis was performed in each patient group. Subdistributional hazard ratios (sHRs) and 95% confidence intervals (95%CIs) were calculated. RESULTS: Overall, 238 patients with 027+ CDI and 267 with 027- CDI were analysed. On multivariate analysis metronidazole monotherapy (sHR 2.380, 95%CI 1.549-3.60, p <0.001) and immunosuppressive treatment (sHR 3.116, 95%CI 1.906-5.090, p <0.001) were factors associated with recurrence in patients with 027+ CDI. In this patient group, metronidazole monotherapy was independently associated with recurrence in both mild/moderate (sHR 1.894, 95%CI 1.051-3.410, p 0.033) and severe CDI (sHR 2.476, 95%CI 1.281-4.790, p 0.007). Conversely, non-severe disease (sHR 3.704, 95%CI 1.437-9.524, p 0.007) and absence of chronic renal failure (sHR 16.129, 95%CI 2.155-125.000, p 0.007) were associated with recurrence in 027- CDI. CONCLUSIONS: Compared to vancomycin, metronidazole monotherapy appears less effective in curing CDI without relapse in the 027+ patient group, independently of disease severity.

A five year retrospective study on Syphilis in the Sexual Transmitted Disease Centre (STDC) of the teaching Hospital Umberto I in Rome.

OBJECTIVES: A retrospective study describing syphilis epidemiological and clinical features in patients referring to an infectious diseases centre in Rome, Italy. METHODS: Between January 2011 and December 2015 demographic, behavioral and clinical data were collected from all adult patients attending the Sexually Transmitted Diseases Centre of the Teaching Hospital Umberto I in Rome. RESULTS: Overall 723 patients, 495 males and 228 females, with syphilis infection diagnosis were included. Average age 39.6 ± 13.6 years (median 38) was higher in men than women (41.1 ± 13.6 vs. 36.3 ± 13.1; p<0.001). Patients were from Italy (486 or 67.2%), EU (90 or 12.4%), rest of Europe (38 or 5.3 %), Americas (46 or 6.4%), Africa (36 or 5.0%) and Asia (27 or 3.7%). One-hundred-twenty-three (17.0%) presented primary syphilis, 43 (5.9%) secondary syphilis, 8 (1.1%) tertiary syphilis, 246 (34.0%) serological syphilis, 80 (11.1%) preceding syphilis, 56 (7.7%) gravidic syphilis and 167 (23.1%) came to the Sexually Transmitted Diseases Centre to control a preceding syphilis treatment. Fifty-six (24.6%) women were diagnosed with syphilis during their pregnancies. Among Chinese female patients, those pregnant represented 87.5%. There were 100 subjects (13.8%) simultaneously HIV+ and 623 (86.2%) HIV- patients. HIV co-infection affected more frequently men (RR 5.30; CI 2.62 - 10.72; p<0.001). In males HIV co-infection affected more frequently homosexuals (RR 11.72; CI 6.72 - 20.45; p<0.001). Overall HIV co-infection affected more frequently foreign patients, specially from the Americas (26.1%), Africa (25.7%) and Asia (22.2%). CONCLUSION: A serious problem of "gravidic syphilis" suggests the need for Public Health preventive action. Also an early diagnosis of both syphilis and HIV infection should be reinforced.

Role of empirical and targeted therapy in hospitalized patients with bloodstream infections caused by ESBL-producing Enterobacteriaceae.

BACKGROUND: Bloodstream infection (BSI) due to extended-spectrum ß-lactamase (ESBL) producing Enterobacteriaceae are a major cause of in-hospital mortality. The effect on survival of empirical and targeted antibiotic therapy in these patients remains controversial. METHODS: A prospective cohort study was conducted analyzing data from 94 patients (age 59 ± 21 years) with BSI due to ESBL producing strains (Sixty-one E. coli, 26 K. pneumoniae, 4 Proteus spp and 3 Enterobacter spp). RESULTS: Risk factors associated with 21-day mortality at univariate analysis were: recent administration of antibiotic therapy (p=0.049), higher SOFA score (p=0.05), ICU stay (p <0.01), hypotension at presentation (p =0.001) or septic shock (p <0.001) and bacteremia from source other than urinary tract (p=0.03). Regardless of antibiotic class used, no differences in survival were noted between patients receiving or not adequate initial antimicrobial treatment (37.1% vs 23.7% p=0.23); on the other hand, compared with the administration of other in vitro active antibiotics, the use of carbapenem as definitive therapy was associated with a significantly lower 21-day mortality (54.3% vs 28.5% p=0.02). CONCLUSIONS: These findings suggest that the administration of an adequate initial therapy is not associated with mortality in hospitalized patients with BSI due to Enterobacteriaceae. The severity of clinical conditions at presentation and the administration of carbapenems as definitive therapy seems to be really important in affecting the outcome of patients with BSI due to ESBL producing strains.

Metal complexes of lactoferrin and their effect on the intracellular multiplication of Legionella pneumophila.

The action of bovine lactoferrin saturated with iron, zinc and manganese on the intracellular multiplication of Legionella pneumophila in HeLa cells has been tested. The results obtained showed that lactoferrin did not influence the invasive efficiency of Legionella. The intracellular multiplication of the bacterium was inhibited by apo-lactoferrin and by lactoferrin saturated with manganese and zinc, whereas lactoferrin saturated with iron enhanced the intracellular growth. Experiments in parallel were performed with iron, manganese and zinc citrate to test the effect due to the metal ions alone. Even in this condition the addition of an iron chelate enhanced the multiplication of Legionella while the manganese chelate produced a certain inhibition.

Multiple types of Legionella pneumophila serogroup 6 in a hospital heated-water system associated with sporadic infections.

Five sporadic cases of nosocomial Legionnaires' disease were documented from 1989 to 1997 in a hospital in northern Italy. Two of them, which occurred in a 75-year-old man suffering from ischemic cardiopathy and in an 8-year-old girl suffering from acute leukemia, had fatal outcomes. Legionella pneumophila serogroup 6 was isolated from both patients and from hot-water samples taken at different sites in the hospital. These facts led us to consider the possibility that a single clone of L. pneumophila serogroup 6 had persisted in the hospital environment for 8 years and had caused sporadic infections. Comparison of clinical and environmental strains by monoclonal subtyping, macrorestriction analysis (MRA), and arbitrarily primed PCR (AP-PCR) showed that the strains were clustered into three different epidemiological types, of which only two types caused infection. An excellent correspondence between the MRA and AP-PCR results was observed, with both techniques having high discriminatory powers. However, it was not possible to differentiate the isolates by means of ribotyping and analysis of rrn operon polymorphism. Environmental strains that antigenically and chromosomally matched the infecting organism were present at the time of infection in hot-water samples taken from the ward where the patients had stayed. Interpretation of the temporal sequence of events on the basis of the typing results for clinical and environmental isolates enabled the identification of the ward where the patients became infected and the modes of transmission of Legionella infection. The long-term persistence in the hot-water system of different clones of L. pneumophila serogroup 6 indicates that repeated heat-based control measures were ineffective in eradicating the organism.

Legionnaires' disease on a cruise ship linked to the water supply system: clinical and public health implications.

The occurrence of legionnaires' disease has been described previously in passengers of cruise ships, but determination of the source has been rare. A 67-year-old, male cigarette smoker with heart disease contracted legionnaires' disease during a cruise in September 1995 and died 9 days after disembarking. Legionella pneumophila serogroup 1 was isolated from the patient's sputum and the ship's water supply. Samples from the air-conditioning system were negative. L. pneumophila serogroup 1 isolates from the water supply matched the patient's isolate, by both monoclonal antibody subtyping and genomic fingerprinting. None of 116 crew members had significant antibody titers to L. pneumophila serogroup 1. One clinically suspected case of legionnaires' disease and one confirmed case were subsequently diagnosed among passengers cruising on the same ship in November 1995 and October 1996, respectively. This is the first documented evidence of the involvement of a water supply system in the transmission of legionella infection on ships. These cases were identified because of the presence of a unique international system of surveillance and collaboration between public health authorities.

Multiplication of Legionella pneumophila in HeLa cells in the presence of cytoskeleton and metabolic inhibitors.

A study has been carried out on the action of cytoskeleton and metabolic inhibitors on intracellular multiplication in HeLa cells of a virulent strain of Legionella pneumophila serogroup 6. The effects of the substances were separately tested on both penetration and intracellular multiplication of L. pneumophila. Only cytochalasin A and 2-deoxy-D-glucose (2dG) affected bacterial internalisation, whereas intracellular multiplication was inhibited by cytochalasins A, B, C, D and J (D being the most active) and by 2dG with a dose-response effect. The action of 2dG was counteracted by 50 mM glucose. Experiments carried out with cytochalasin D and a rhodamine-phalloidin conjugate showed the involvement of cytoskeletal elements in intracellular multiplication of Legionella; compounds acting on microtubules had no effect.

Bafilomycin A1 and intracellular multiplication of Legionella pneumophila.

Multiplication of Legionella pneumophila in HeLa cells was found to be inhibited by noncytotoxic concentrations of bafilomycin A1, with blockage of bacterial growth at a concentration 15.6 nM. The inhibiting action was evident only when the antibiotic was present during the initial phase of intracellular multiplication, i.e., during the formation of the phagosome, whereas the addition of the drug did not affect microorganisms already actively multiplying within the phagosome.

Molecular epidemiology of an outbreak of Legionnaires' disease associated with a cooling tower in Genova-Sestri Ponente, Italy.

Fatty acid profile analysis, monoclonal antibody (MAb) subtyping, pulsed-field gel electrophoresis (PFGE), arbitrarily primed polymerase chain reaction (AP-PCR), and ribotyping were used to compare clinical and environmental Legionella pneumophila serogroup 1 isolates from an outbreak of Legionnaires' disease presumptively associated with cooling towers. According to the Oxford subtyping scheme, the MAb subtype of patients' isolates and of two strains originating from a cooling tower was Pontiac, whereas the other isolates were subtype Olda. The strains showed no intrinsic strain-to-strain difference in fatty acid profiles, and ribotyping and length polymorphism of the 16S-23S rDNA intervening regions failed to reveal any differences between the isolates. Conversely, PFGE and AP-PCR appeared to be more discriminatory, as the same genomic profile was found for the clinical and some environmental strains. Meteorologic and epidemiological data and the results of molecular analysis of the Legionella pneumophila serogroup 1 isolates support the hypothesis that the infection was transmitted from one of the cooling towers to the indoor environment of the same building, to homes in proximity that had open windows, and to the streets. In fact, the outbreak diminished and later ended after a part in the tower was replaced. This investigation demonstrates the utility of combined molecular methods (i.e., phenotypic and genomic typing) in comparing epidemiologically linked clinical and environmental isolates. Finally, the outbreak confirms the risk of Legionnaires' disease posed by cooling towers, mainly when atmospheric thermal and humidity inversions occur. This finding emphasizes the need to determine whether the source of infection is in the living or working environment or somewhere else.

Effect of monensin on the invasiveness and multiplication of Legionella pneumophila.

The polyether antibiotic monensin exhibited bacteriostatic activity against a clinical isolate of Legionella pneumophila in vitro. Experiments designed to test the effect of the compound on the invasiveness and multiplication of L. pneumophila in HeLa cells showed that, in the presence of the antibiotic, legionellas that penetrated the cells did not multiply. However, monensin did not alter the characteristics of phagosomes that contained ingested legionellas. In the presence of monensin, infected cells exhibited extensive vacuolation and a noticeable reduction in the number of intracellular micro-organisms was evident a few hours after infection.

Effect of various detergents on the interaction between BK virus and susceptible cells.

The activity of various types of detergents towards the infectivity and the hemagglutinating activity of BK virus was studied. Similarly to other non enveloped small viruses, BK virus showed a significant resistance to non cytotoxic concentrations of the compounds tested, with the exception of N-Dodecyl-N,N-dimethyl-3-ammonio-1-propane-sulfonate. In some cases, mainly with N-Octylglucoside, both the infectivity and the hemagglutinating activity of BK virus was enhanced.

Effect of sub-inhibitory concentrations of antibiotics on the hemolytic activity of Legionella.

The effect of sub-inhibitory concentrations of various antibiotics on the hemolytic activity of different strains of Legionella has been tested. By means of a gradient plate technique it was possible to demonstrate that in a limited range of sub-inhibitory concentrations, antibiotics did not affect bacterial growth but inhibited the hemolytic activity of the strains examined.

Role of phospholipids in BK virus infection and haemagglutination.

The role of phospholipids in BK virus infection and haemagglutination was studied by competition binding experiments and by treatment of susceptible cells with phospholipases. Phospholipids extracted from Vero cells and some commercial phospholipids showed an inhibiting activity on both BK virus infectivity and haemagglutination. The treatment of Vero cells with phospholipases affected the binding of BK virus, but the addition of phospholipids to enzyme-treated cells restored their susceptibility to both viral infectivity and haemagglutination.

Effect of biological and synthetic polymers on BK virus infectivity and hemagglutination.

The effect of several biological and synthetic polymers, chosen on the basis of different physical and chemical properties, was investigated on BK virus infectivity and hemagglutination. It was observed that polyanions like mucin, dextran sulfate and heparin depressed the viral binding, whereas polycations had no significant activity, with the exception of poly-L-lysine, which enhanced it. The effect of the active polymers was studied in different experimental conditions and the results obtained suggested that polyanions may act directly on the virus particle, whereas the target of polycations could be at the level of cell membranes. However, the effect shown by the active compounds did not appear to be simply related to the electric charge since neutral compounds, such as tamarind gum and locust bean gum, showed a marked inhibitory effect on BK virus binding to the cells.

Growth of Legionella spp. under conditions of iron restriction.

The growth inhibiting activity of transferrins, citrate, 2-2' dipyridyl and desferrioxamine methanesulphonate towards Legionella spp. and their serogroups was investigated. The inhibitory activity of all these compounds depended upon the iron-free state of the molecules and was abolished by saturation with iron. No bactericidal effect by transferrins was observed at concentrations up to four times the minimal bacteriostatic concentration. No interaction of transferrins with the legionella cell surface was detected by direct or indirect fluorescence assay, or by dialysis culture experiments in which transferrin was separated from the bacterial cells. The demonstration of a siderophore-like activity in supernates of iron-deficient legionella cultures may account for the ability of Legionella spp. to multiply in conditions of iron restriction.

Extraction and purification of gangliosides from CER cells, a cell line suitable for rabies virus replication.

Results are reported concerning the extraction and purification of gangliosides present in CER cells, a cell line which is highly suitable for the cultivation in vitro of rabies virus. By means of thin layer chromatography it has been possible to demonstrate that the CER cell ganglioside pattern is composed mainly of monosialo and disialogangliosides.

Legionella pneumophila serogroup 1 population in Italy by monoclonal subtyping.

The Oxford panel of monoclonal antibodies was used to subtype 83 strains of Legionella pneumophila serogroup 1 of human and environmental origin. The International panel was also used to subtype 50 of them. All the 18 patients' isolates were of the Pontiac subgroup, and 40/43 of the environmental strains of the Pontiac subgroup were associated with human infection. The remaining environmental strains were subgroups Olda (15 strains), Camperdown (5 strains), and Bellingham (2 strains). The Philadelphia subgroup was the commonest among the environmental strains tested with the international MABs panel. This study confirms previous findings that L. pneumophila serogroup 1 isolates with the Pontiac (Oxford panel) or MAB-2 (international panel) reacting antigen marker seem to be more virulent than the other subgroups.

Involvement of gangliosides in the interaction between BK virus and Vero cells.

BK virus infectivity was inhibited by gangliosides extracted from Vero cells and by standard preparations of different gangliosides. Gangliosides were also able to restore the susceptibility of glycosidase-treated Vero cells to BK virus infection.