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Background A wide range of frequency of azole-resistance in A fumigatus in different patient populations worldwide was observed threatening to reduce therapeutic options. Objectives Estimate the prevalence of azole-resistance, investigate the molecular mechanisms of resistance, compare the genotypes of resistant clinical isolates with those from the surrounding environment. Methods Aspergillus isolates were collected by seven Italian hospital microbiology laboratories. Strains were isolated from different clinical samples from unselected patients. The azole-resistance was evaluated using screening test and microdilution EUCAST method. The molecular mechanism of resistance was performed sequencing the cyp51A gene. Resistant isolates were genotyped by microsatellite analysis and their profiles compared with those of azole-resistant isolates from previous Italian studies. Results 425 Aspergillus isolates from 367 patients were analysed. The azole-resistance rates were 4.9% and 6.6% considering all Aspergillus spp. isolates and the A fumigatus sensu stricto, respectively. All resistant isolates except one were from a single hospital. Two rare azole-resistant species were identified: A thermomutatus and A lentulus. The predominant resistance mechanism was TR34 /L98H. No correlation between the clinical resistant strains and environmental isolates from patients' home/work/ward was observed. The analysis of the molecular correlation between the resistant clinical strains collected in the present study and those of environmental and clinical origin collected in previous Italian studies reveals a progressive diversification of azole-resistant genotypes starting from a founder azole-resistant genotype. Conclusions This study confirms the trend of azole-resistance rate in Italy, showing a geographical difference. Data reinforce the importance of surveillance programmes to monitor the local epidemiological situation.

Objectives This study was conducted to assess the prevalence of azole resistance in Aspergillus isolates from patients with haematological malignancies or who were undergoing haematopoietic stem cell transplantation and to identify the molecular mechanism of resistance. Methods In this 28-month prospective study involving 18 Italian centres, Aspergillus isolates from surveillance cultures were collected and screened for azole resistance, and mutations in the cyp51A gene were identified. Resistant isolates were genotyped by microsatellite analysis, and the allelic profiles were compared with those of resistant environmental and clinical isolates from the same geographical area that had been previously genotyped. Results There were 292 Aspergillus isolates collected from 228 patients. The isolates belonged mainly to the section Fumigati (45.9%), Nigri (20.9%), Flavi (16.8%) and Terrei (4.8%). Three isolates showed itraconazole resistance: Aspergillus fumigatus sensu stricto, Aspergillus lentulus (section Fumigati) and Aspergillus awamori (section Nigri). The itraconazole resistance rates were 1% and 1.48% considering all Aspergillus spp. isolates and the Aspergillus section Fumigati, respectively. The prevalence of azole resistance among all the patients was 1.3%. Among patients harbouring A. fumigatus sensu stricto isolates, the resistance rate was 0.79%. The A. fumigatus isolate, with the TR34/L98H mutation, was genotypically distant from the environmental and clinical strains previously genotyped. Conclusions In this study, the Aspergillus azole resistance rate was 1% (3/292). In addition to A. fumigatus sensu stricto, A. lentulus and A. awamori azole-resistant isolates were identified. Therefore, it is important have a correct identification at the species level to address a rapid therapy better, quickly understand the shift towards cryptic species and have an updated knowledge of the local epidemiology.

Analisi delle prospettive di sviluppo del traffico marittimo in project cargo e dei punti di forza e debolezza del porto di Livorno

In 2012 we undertook a screening study for Enterobacteriaceae with extended-spectrum β-lactamases (ESBLs), derepressed or acquired high-level AmpC cephalosporinases, and metallo-β-lactamases (MBLs), and also methicillin-resistant Staphylococcus aureus (MRSA), in a long-term care facility (LTCF1) and the associated acute care hospital geriatric ward in Bolzano, northern Italy. The study followed up an initial survey carried out in LTCF1 in 2008. For comparison, screening in 2012 was extended to a second LTCF.Urine samples and rectal, inguinal, oropharyngeal, and nasal swabs were plated on selective agars. Isolates were typed by pulsed-field gel electrophoresis. Resistance genes and Escherichia coli belonging to ST131 were sought by PCR. Demographic data were collected.Fewer residents of LTCF1 were colonized with multidrug-resistant (MDR) bacteria in 2012: all MDR organisms, 53.8% vs 74.8% in 2008; ESBL producers, 49.0% vs 64.0% in 2008; MRSA, 13.2% vs 38.7% in 2008; only 2 MBL-producers were isolated in 2012 vs 8 in 2008. Colonization of staff in LTCF1 by MDR bacteria had also decreased (overall 10.5% in 2012 vs 27.5% in 2008). Changed case mixes and risk factors, together with strengthened hygiene measures probably underlie the changes. Colonization proportions in 2012 in LTCF2 were similar to those in LTCF1. By contrast there was no significant change in the proportion of patients colonized by MDR bacteria in the geriatric ward (22.2% in 2008 vs 22.7% in 2012).A significant decrease in the proportions of staff and residents of an LTCF colonized by MDR bacteria was observed over a 4-y interval.

Panton–Valentine leukocidin (PVL)-positive community-associated meticillin-resistant Staphylococcus aureus (CA-MRSA) isolates are widespread in many countries, with varying distribution and epidemiology. The aim of this study was to characterise 10 PVL-positive MRSA isolates collected during February 2010 to January 2011 from skin and soft-tissue infections in the North Italian Province of Bolzano. Accessory gene regulator ( agr ) typing, staphylococcal cassette chromosome mec (SCC mec ) typing, staphylococcal protein A ( spa ) gene typing, multilocus sequence typing, toxin gene profiling, polymerase chain reaction for type I arginine catabolic mobile element (ACME) and antimicrobial resistance typing were applied to the isolates. Eight different CA-MRSA clones were identified, including ST30-IVc, ST772-V, ST80-IVc, ST5-IVc, ST88-IVa, ST93-IVa, ST8-IVc and the type I ACME-positive ST8-IVa. The high heterogeneity of PVL-positive MRSA probably reflects the introduction of different clones by international travellers or immigrants.

Capnocytophaga canimorsus is a commensal bacterium in the oral cavity of dogs and cats and has been isolated from humans with a history of dog or cat bites or scratches. C. canimorsus are capnophilic facultative anaerobic, fastidious gram negative fusiform rods, included in the CDC group DF-2 or dysgonic fermenter 2. The bacterium can cause severe illness in patients at elevated risk i.e. after splenectomy, with alcohol addiction, chronic lung or liver diseases or immunosuppression. In 2010 at the Bolzano Regional Hospital (Northern-Italy) C. canimorsus was isolated from blood cultures in two splenectomised patients suffering from septicaemia.The bacterium is difficult to cultivate because of its specific requirements for nutrients as well as to identify with routine automated instruments; therefore we used molecular typing by 16S rDNA sequencing.The true number of C. canimorsus infections is probably underestimated due to the fastidious growth of the organism. However, given the severity of the disease and the poor outcome one should consider this bacterium to apply appropriate diagnostic methods for this etiologic agent.

Paecilomyces lilacinus, a filamentous fungus living saprophytic in the environment, very rarely causes infection in immunocompetent humans, mostly ocular or cutaneous and sub-cutaneous infections.The fungal isolates usually show low susceptibility to conventional antifungal drugs in vitro and variable susceptibility to novel triazoles. During a period of 2 years, 4 cultures of nail fragments were grown from a 41 years old female on modified Sabouraud/Dextrose agar (S.D.A.) (CAF bioMerieux, CAF+CEX Biolife). Cultures were continued in suspension and the hyphae stained with lactophenole blue analysed microscopically. 2 samples were tested with molecular methods, i.e. DNA extraction, amplification of internal transcribed spacer (ITS) region 1 and 2 and automated sequencing. The sequence results were evaluated using a validated reference database for humanpathogenic fungi. Additionally the BLAST algorithm on NIH database GenBank was used. A growth of a filamentous hyaline fungus with white colonies and lila shade was observed in all samples. The presence of Penicillium like conidiophores with divergent long terminal phialides was seen with little elliptical conidia. DNA sequencing determined the species as Paecilomyces lilacinus with 99% homology with GenBank isolate AY213667. The infection of the right hallux did not improve by systemic (Lamisil/terbinafine) and local treatment (Locetar/amorolfin nail lacquer).Also change to Sporanox/itraconazol only led to deterioriation, so finally the extraction of the nail had to be performed given the known difficulties with treatment described in the literature. This case highlights the usefulness of molecular techniques, especially use of the highly discriminative ITS region for fungal typing.