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Objectives To detect possible severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) RNA contamination of inanimate surfaces in areas at high risk of aerosol formation by patients with coronavirus disease 2019 (COVID-19). Methods Sampling was performed in the emergency unit and the sub-intensive care ward. SARS-CoV-2 RNA was extracted from swabbed surfaces and objects and subjected to real-time RT-PCR targeting RNA-dependent RNA polymerase and E genes. Virus isolation from positive samples was attempted in vitro on Vero E6 cells. Results Twenty-six samples were collected and only two were positive for low-level SARS-CoV-2 RNA, both collected on the external surface of continuous positive airway pressure helmets. All transport media were inoculated onto susceptible cells, but none induced a cytopathic effect on day 7 of culture. Conclusions Even though daily contact with inanimate surfaces and patient fomites in contaminated areas may be a medium of infection, our data obtained in real-life conditions suggest that it might be less extensive than hitherto recognized.

Objective: This study aimed to assess the role of Tocilizumab therapy (TCZ) in terms of ICU admission and mortality rate of critically ill patients with severe COVID-19 pneumonia. Design: Patients with COVID-19 pneumonia were prospectively enrolled in SMAtteo COvid19 REgistry (SMACORE). A retrospective analysis of patients treated with TCZ matched using propensity score to patients treated with Standard Of Care (SOC) was conducted. Setting: The study was conducted at IRCCS Policlinico San Matteo Hospital, Pavia, Italy, from March 14, 2020 to March 27, 2020. Participants: Patients with a confirmed diagnosis of COVID-19 hospitalized in our institution at the time of TCZ availability. Interventions: TCZ was administered to 21 patients. The first administration was 8 mg/kg (up to a maximum 800 mg per dose) of Tocilizumab intravenously, repeated after 12 h if no side effects were reported after the first dose. Main Outcomes and Measures: ICU admission and 7-day mortality rate. Secondary outcomes included clinical and laboratory data. Results: There were 112 patients evaluated (82 were male and 30 were female, with a median age of 63.55 years). Using propensity scores, the 21 patients who received TCZ were matched to 21 patients who received SOC (a combination of hydroxychloroquine, azithromycin and prophylactic dose of low weight heparin). No adverse event was detected following TCZ administration. This study found that treatment with TCZ did not significantly affect ICU admission (OR 0.11; 95% CI between 0.00 and 3.38; p = 0.22) or 7-day mortality rate (OR 0.78; 95% CI between 0.06 and 9.34; p = 0.84) when compared with SOC. Analysis of laboratory measures showed significant interactions between time and treatment regarding C-Reactive Protein (CRP), alanine aminotransferase (ALT), platelets and international normalized ratio (INR) levels. Variation in lymphocytes count was observed over time, irrespective of treatment. Conclusions: TCZ administration did not reduce ICU admission or mortality rate in a cohort of 21 patients. Additional data are needed to understand the effect(s) of TCZ in treating patients diagnosed with COVID-19.

The activity of voriconazole was tested in vitro against 1996 clinical yeast isolates collected in 20 Italian microbiology laboratories. Voriconazole susceptibility testing was carried out with the broth microdilution (NCCLS M27-A2), Etest and disk diffusion methods. The minimum inhibitory concentrations at which 90% of the isolates were inhibited (MIC90) obtained with the NCCLS method were 0.03 mg/L for Candida albicans, 0.5 mg/L for Candida non-albicans and 0.25 mg/L for other genera; those obtained with Etesting were, respectively, 0.032 mg/L, 0.125 mg/L and 0.125 mg/L. With the disk diffusion method, the majority of isolates (92.3%) showed inhibition zone diameters between 21 mm and 40 mm. Using a tentative MIC cut-off of 1 mg/L as indicative of in vitro susceptibility, 98.1% of the isolates tested in our study would be classified as susceptible, and only 28 (1.4%) of the isolates, with MICs higher than 2 mg/L, would be classified as resistant to the drug. Our findings confirm the broad-spectrum in vitro activity of voriconazole against yeasts, including Candida species that are generally less susceptible to other azoles.

We evaluated the in vitro activity of voriconazole, amphotericin B, and itraconazole against 192 clinical mould isolates recovered in twenty Italian microbiology laboratories. The vast majority of isolates belonged to the genus Aspergillus (94.2%) with A. fumigatus (58.3%) being the most frequently isolated species. Antifungal susceptibility testing was performed using the broth microdilution method defined by the CLSI M38-A standard, and results were compared to those obtained with Sensititre panels. Aspergillus flavus ATCC 204304 was employed as reference strain and results were within all expected ranges. Voriconazole's activity against the 192 mould isolates was comparable to that of amphotericin B and itraconazole: voriconazole MIC90 (CLSI 1 microg/ml, Sensititre 1 microg/ml), itraconazole MIC90 (CLSI 0.5 microg/ml, Sensititre 0.5 microg/ml), amphotericin B MIC90 (CLSI 1 microg/ml, Sensititre 1 microg/ml). In conclusion, these in vitro data highlight voriconazole's broad spectrum activity against filamentous fungi and support its use as a first line agent for the treatment of fungal diseases.

Midichloria mitochondrii is an intracellular bacterium found in the hard tick Ixodes ricinus. In this arthropod, M. mitochondrii is observed in the oocytes and in other cells of the ovary, where the symbiont is present in the cell cytoplasm and inside the mitochondria. No studies have so far investigated whether M. mitochondrii is present in the salivary glands of the tick and whether it is transmitted to vertebrates during the tick blood meal. To address the above issues, we developed a recombinant antigen of M. mitochondrii (to screen human sera) and antibodies against this antigen (for the staining of the symbiont). Using these reagents we show that (i) M. mitochondrii is present in the salivary glands of I. ricinus and that (ii) seropositivity against M. mitochondrii is highly prevalent in humans parasitized by I. ricinus (58%), while it is very low in healthy individuals (1·2%). These results provide evidence that M. mitochondrii is released with the tick saliva and raise the possibility that M. mitochondrii is infectious to vertebrates. Besides this, our study indicates that M. mitochondrii should be regarded as a package of antigens inoculated into the human host during the tick bite. This implies that the immunology of the response toward the saliva of I. ricinus is to be reconsidered on the basis of potential effects of M. mitochondrii and poses the basis for the development of novel markers for investigating the exposure of humans and animals to this tick species.

Sequence type 258 (ST258) is the most widespread multidrug resistant (MDR) Klebsiella pneumoniae strain worldwide. Here, we report the draft genome sequences of two colistin-resistant MDR K. pneumoniae ST258 clinical strains isolated from hospital patients in Italy. These strains are resistant to β-lactams, cephalosporins, fluoroquinolones, aminoglycosides, macrolides, tetracyclines, carbapenems, and colistin.

Here, we present the draft genome sequence of Klebsiella pneumoniae subsp. pneumoniae sequence type 512 (ST512) isolated during a KPC-producer outbreak. This strain is resistant to β-lactams, cephalosporins, fluoroquinolones, aminoglycosides, macrolides, tetracyclines, and carbapenems but susceptible to colistin. The ST512-K30BO genome is composed of 289 contigs for 5,392,844 bp with 56.9% G+C content.

a b s t r a c t Background: Specific bacterial infections or alterations of the gut microbiota likely trigger immuno- pathological phenomena associated with Crohn's disease and ulcerative colitis. Mycobacterium avium subspecies paratuberculosis is a candidate etiological agent of Crohn's disease. Definitive causal connec- tion between Mycobacterium avium subspecies paratuberculosis infection and Crohn's disease has not been demonstrated. Aims: To determine the circulation of Mycobacterium avium subspecies paratuberculosis in Crohn's disease patients and water supplies in an Italian region where this bacterium is endemic in cattle farms. Methods: Mycobacterium avium subspecies paratuberculosis screening was performed on biopsies from human patients, and from water samples, using two different PCR procedures. Results: In hospitals where multiple specimens were obtained from different sites in the intestine, the prevalence of Mycobacterium avium subspecies paratuberculosis infection was 82.1% and 40% respec- tively in Crohn's disease and ulcerative colitis patients; in another hospital, where single specimens were obtained from patients, the bacterium was not detected. Control subjects also harboured Mycobac- terium avium subspecies paratuberculosis, but at a lower prevalence. Tap water samples collected in the study area contained Mycobacterium avium subspecies paratuberculosis DNA. Discussion: The results of screenings for Mycobacterium avium subspecies paratuberculosis in humans are deeply influenced by both the number and location of the collected biopsies. There is a wide circulation of the organism in the study area, considering the prevalence in humans and its presence in drinking water.

To confirm thatMycobacterium tuberculosischaperonin 10 (Cpn10) is secreted outside the live bacillus, infected macrophages were examined by electron microscopy. This revealed that the mycobacterial protein accumulates both in the wall of the bacterium and in the matrix of the phagosomes in which ingested mycobacteria survive within infected macrophages. To understand the structural implications underlying this secretion, a structural study ofM.tuberculosisCpn10 was performed under conditions that are generally believed to mimic the membrane environment. It was found that in buffer-organic solvent mixtures, the mycobacterial protein forms two main species, namely, a partially helical monomer that prevails in dilute solutions at room temperature and a dimer that folds into a β-sheet-dominated structure and prevails in either concentrated protein solutions at room temperature or in dilute solutions at low temperature. A partially helical monomer was also found and was completely associated with negatively charged detergents in a micelle-bound state. Remarkably, zwitterionic lipids had no effect on the protein structure. By using N- and C-truncated forms of the protein, the C- and N-terminal sequences were identified as possessing an amphiphilic helical character and as selectively associating with acidic detergent micelles. When the study was extended to other chaperonins, it was found that human Cpn10 is also monomeric and partially helical in dilute organic solvent-buffer mixtures. In contrast,Escherichia coliCpn10 is mostly dimeric and predominately β-sheet in both dilute and concentrated solutions. Interestingly, human Cpn10 also crosses biological membranes, whereas theE.colihomologue is strictly cytosolic. These results suggest that dissociation to partially helical monomers and interaction with acidic lipids may be two important steps in the mechanism of secretion ofM.tuberculosisCpn10 to the external environment.