Four-Year Environmental Surveillance Program of Legionella spp. in One of Palermo’s Largest Hospitals

Legionella is a ubiquitous bacterium that lives in freshwater environments and colonizes human-made water systems. Legionella pneumophila is the most virulent species, and risk factors for Legionnaires’ disease include increasing age, smoking, chronic diseases, and immunodeficiency. For this reason, it is very important to assess and monitor hospital water systems in order to prevent legionellosis. We have monitored a large hospital in Palermo for four years. To determine the presence of microorganisms, according to national guidelines, we used the culture method, which is considered the gold standard for Legionella detection. Sampling was divided into five macro-areas, and a total of 251 samples were collected during the period of investigation, 49% of which were Legionella spp.- positive and 51% were Legionella spp.-negative. Positive samples with L. pneumophila. sgr 2-15 were most frequent in the Underground (55.6%, p = 0.0184), Medicine (42.9%, p = 0.0184) and Other (63.2%, p = 0.002) areas; while positive samples for L. pneumophila sgr 1 were less frequent in the Underground (0.0%, p = 0.0184) and Surgery areas (4.5%, p = 0.033), and for Legionella anisa, were less frequent in the Medicine (4.1%, p = 0.021), Oncohematology (0.0%, p = 0.0282), and Other (0.0%, p = 0.016) areas. Finally, no significant differences were observed among the areas for each isolate considered. The surveillance carried out in these years demonstrates the importance of monitoring, which allows us to analyze the conditions of hospital facilities and, therefore, prevent Legionella spp. infections

Multicomponent Antibiofilm Lipid Nanoparticles as Novel Platform to Ameliorate Resveratrol Properties: Preliminary Outcomes on Fibroblast Proliferation and Migration

: The well-being of skin and mucous membranes is fundamental for the homeostasis of the body and thus it is imperative to treat any lesion quickly and correctly. In this view, polyphenols might assist and enhance a successful wound healing process by reducing the inflammatory cascade and the production of free radicals. However, they suffer from disadvantageous physico-chemical properties, leading to restricted clinical use. In this work, a complex mixture of PEGylated lipid, Glyceryl monoester, 18-β-Glycyrrhetinic Acid and Menthol was designed to entrap Resveratrol (RSV) as the active ingredient and further produce lipid nanoparticles (LNPs) by homogenization followed by high-frequency sonication. The nanosystem was properly characterized in terms of particle size (DLS, SEM), zeta potential, drug loading, antioxidant power (DPPH), release behaviour, cytocompatibility, wound healing and antibiofilm properties. The optimized lipid mixture was homogeneous, melted at 57-61 °C and encapsulated amorphous RSV (4.56 ± 0.04% w/w). The RSV-loaded LNPs were almost monodispersed (PDI: 0.267 ± 0.010), with nanometric size (162.86 ± 3.12 nm), scavenger properties and suitable DR% and LE% values (96.82 ± 1.34% and 95.17 ± 0.25%, respectively). The release studies were performed to simulate the wound conditions: 1-octanol to mimic the lipophilic domains of biological tissues (where the First Order kinetic was observed) and citrate buffer pH 5.5 according to the inflammatory wound exudate (where the Korsmeyer-Peppas kinetic was followed). The biological and microbiological evaluations highlighted fibroblast proliferation and migration effects as well as antibiofilm properties at extremely low doses (LNPs: 22 μg/mL, corresponding to RSV 5 µM). Thus, the proposed multicomponent LNPs could represent a valuable RSV delivery platform for wound healing purposes

Multicentre Surveillance of Candida Species from Blood Cultures during the SARS-CoV-2 Pandemic in Southern Europe (CANCoVEU Project)

Introduction: Surveillance of Candida species isolates from blood cultures (BCs) in Europe is considered fragmented, unable to allow the definition of targets of antifungal stewardship recommendations especially during the SARS-CoV-2 pandemic. Methods: We performed a multicentric retrospective study including all consecutive BC Candida isolates from six Southern European tertiary hospitals (1st January 2020 to 31st December 2021). Etiology, antifungal susceptibility patterns, and clinical setting were analyzed and compared. Results: C. albicans was the dominant species (45.1%), while C. auris was undetected. Candida species positive BC events increased significantly in COVID-19 ICUs in 2021 but decreased in other ICUs. Resistance to azole increased significantly and remained very high in C. albicans (fluconazole from 0.7% to 4.5%, p = 0.03) and C. parapsilosis complex (fluconazole up to 24.5% and voriconazole up to 8.9%), respectively. Resistance to caspofungin was remarkable in C. tropicalis (10%) and C. krusei (20%), while resistance to at least one echinocandin increased in 2021, especially in C. parapsilosis complex (from 0.8% to 5.1%, p = 0.05). Although no significant differences were observed over the study period, fluconazole and echinocandin resistance increased in COVID-19 ICUs by up to 14% and 5.8%, respectively, but remained undetected in non-intensive COVID-19 wards. Conclusions: Antifungal stewardship activities aimed at monitoring resistance to echinocandin in C. tropicalis and C. krusei, and against the spread of fluconazole resistant C. parapsilosis complex isolates are highly desirable. In COVID-19 patients, antifungal resistance was mostly present when the illness had a critical course.publishersversionpublishe

Improvement of a rapid direct blood culture microbial identification protocol using MALDI-TOF MS and performance comparison with SepsiTyper kit

Fast diagnosis of pathogens is critical to guarantee the most adequate therapy for infections; bacterial culture methods, which constitute the actual gold standard, are precise and sensitive but rather slow. Today, new methods have been made available to enable faster diagnosis, with the Matrix-Assisted Laser Desorption Ionization\u2013Time Of Flight Mass Spectrometry (MALDI-TOF MS) technique being the most promising. Even if simpler and faster than traditional bacterial culture methods, analysis of positive blood cultures via MALDI-TOF MS requires a preliminary extraction process of samples. In this study, we compared two extraction protocols for bacterial identification directly from positive blood cultures using the Bruker MALDI Biotyper system (Bruker Daltonics, Billerica, MA). In particular, we evaluated the time employed and the overall performance for their accurate identification. In this work, the performances of a commercial extraction kit, named SepsiTyper\u2122 Kit, and those of the protocol developed by Treibmann et al. were evaluated and proven to be similar. However, the SELTERS method represents the best compromise price/performance. Lastly, an in-house developed analysis protocol has been tested, and the introduced optimizations granted a performance level equal if not better than the SepsiTyper kit, a reduced processing time and reduced costs

Transfusion-Transmitted Malaria of Plasmodium malariae in Palermo, Sicily

Transfusion-transmitted malaria (TTM) is a rare occurrence with serious consequences for the recipient. In non-endemic areas, the incidence of transmission of malaria by transfusion is very low. We report a clinical case of transfusion-transmitted malaria due to Plasmodium malariae, which happened in a patient with acute hemorrhagic gastropathy. Case presentation: In April 2019, a 70-year-old Italian man with recurrent spiking fever for four days was diagnosed with a P. malariae infection, as confirmed using microscopy and real-time PCR. The patient had never been abroad, but about two months before, he had received a red blood cell transfusion for anemia. Regarding the donor, we revealed that they were a missionary priest who often went to tropical regions. Plasmodium spp. PCR was also used on donor blood to confirm the causal link. Discussion and Conclusions: The donations of asymptomatic blood donors who are predominantly “semi-immune” with very low parasitic loads are an issue. The main problem is related to transfusion-transmitted malaria. Our case suggests that P. malariae infections in semi-immune asymptomatic donors are a threat to transfusion safety. Currently, microscopy is considered the gold standard for the diagnosis of malaria but has limited sensitivity to detect low levels of parasitemia. Screening using serological tests and molecular tests, combined with the donor’s questionnaire, should be used to reduce the cases of TTM

Four-Year Environmental Surveillance Program of Legionella spp. in One of Palermo’s Largest Hospitals

Legionella is a ubiquitous bacterium that lives in freshwater environments and colonizes human-made water systems. Legionella pneumophila is the most virulent species, and risk factors for Legionnaires’ disease include increasing age, smoking, chronic diseases, and immunodeficiency. For this reason, it is very important to assess and monitor hospital water systems in order to prevent legionellosis. We have monitored a large hospital in Palermo for four years. To determine the presence of microorganisms, according to national guidelines, we used the culture method, which is considered the gold standard for Legionella detection. Sampling was divided into five macro-areas, and a total of 251 samples were collected during the period of investigation, 49% of which were Legionella spp.-positive and 51% were Legionella spp.-negative. Positive samples with L. pneumophila. sgr 2-15 were most frequent in the Underground (55.6%, p = 0.0184), Medicine (42.9%, p = 0.0184) and Other (63.2%, p = 0.002) areas; while positive samples for L. pneumophila sgr 1 were less frequent in the Underground (0.0%, p = 0.0184) and Surgery areas (4.5%, p = 0.033), and for Legionella anisa, were less frequent in the Medicine (4.1%, p = 0.021), Oncohematology (0.0%, p = 0.0282), and Other (0.0%, p = 0.016) areas. Finally, no significant differences were observed among the areas for each isolate considered. The surveillance carried out in these years demonstrates the importance of monitoring, which allows us to analyze the conditions of hospital facilities and, therefore, prevent Legionella spp. infections

Draft Genome Sequence and Biofilm Production of a Carbapenemase-Producing Klebsiella pneumoniae (KpR405) Sequence Type 405 Strain Isolated in Italy

Rapid identification and characterization of multidrug-resistant Klebsiella pneumoniae strains is essential to diagnose severe infections in patients. In clinical routine practice, K. pneumoniae is frequently identified and characterized for outbreak investigation. Pulsed-field gel electrophoresis or multilocus sequence typing could be used, but, unfortunately, these methods are time-consuming, laborious, expensive, and do not provide any information about the presence of resistance and virulence genes. In recent years, the decreasing cost of next-generation sequencing and its easy use have led to it being considered a useful method, not only for outbreak surveillance but also for rapid identification and evaluation, in a single step, of virulence factors and resistance genes. Carbapenem-resistant strains of K. pneumoniae have become endemic in Italy, and in these strains the ability to form biofilms, communities of bacteria fixed in an extracellular matrix, can defend the pathogen from the host immune response as well as from antibiotics, improving its persistence in epithelial tissues and on medical device surfaces

Molecular characterization of Listeria monocytogenes strains isolated from food and human sources

Introduction: The aim of the work was the molecular characterization of L. monocytogenes (L.m.) isolated from food and humans in the period 2019-2021. Materials and methods: A total of 67 L.m. isolates have been collected (UNI EN ISO 11290-1: 2017) at the Food Microbiology laboratories of the IZS Palermo from different food matrices with a prevalence of ready-to-eat food products such as dairy, fresh cheese, meat, fish sampled by Veterinary Services. The bacterial isolates were stored at -20 ° C in Microbank and sent to the LNR of Teramo for the determination of the serogroup (PCR-Multiplex) and molecular investigation (MLST and Whole Genome Sequencing -WGS). The sequencing was carried out on the Illumina NextSeq 500 platform and the data analysis was performed on a bioinformatics and data collection platform of the LNR L.m, called GenPat. N. 39 clinical strains of L.m. were collected at Palermo hospital laboratories (Regional Reference Laboratory for listeriosis). The strains, together with the sheets for the collection of epidemiological data, were sent to the Istituto Superiore di Sanità (ISS- Operational Microbiological Contact Point for L.m.) for molecular characterization by WGS. The sequencing was carried out on an IonTorrent S5 platform and the analysis was performed automatically on a bioinformatics and data collection platform, called IRIDA-ARIES. Results: The results on the molecular characterization of the strains isolated from food showed a prevalence of serogroup IVb (70%), followed by IIa (24%), IIb (4.5%) and IIc (1.5%). The 46.3% of the strains, all belonging to serogroup IVb, have been isolated from milk products (mozzarella and string cheese) taken from the same producer during official controls, in the period August- September 2020, following a positivity detected in self-control. The results of the sequencing carried out by the LNR highlighted the existence of clusters of strains belonging to Clonal Complex (CC) 2, CC199 and CC1. In particular, all the strains isolated in the same cheese factory, both in the period considered and in a subsequent sampling in January 2022, belonged to CC2 and Sequence Type 2 (ST2). Regarding the 39 clinical L.m. strains, the phylogenetic analysis of the genomes allowed the identification of a Cluster_90 composed of 25 strains of which 6 were isolated in 2019, 16 in 2020 and 1 in 2021. The 25 strains were found to belong to serogroup IVb, ST2 and CC2, presenting between them an allelic difference in the MLST core genome between 0 and 12. It has been noted the presence of 8 strains, isolated between October 2019 and August 2021, associated with maternal and neonatal cases. The genomic sequences of the 39 clinical strains of L.m. were compared with the sequences deposited in the IRIDA-ARIES database. From the analysis, it emerged that the Cluster_90, ST2 also included 3 isolated strains in Lombardy (2 in 2019, 1 in 2020), 2 in Piedmont, 1 in Lazio, 1 in Tuscany in 2020 and 2 isolated strains in Sicily (1 in 2019 from Syracuse and 1 in 2020 from the province of Palermo). Conclusion: This study allowed to start the construction of an integrated medical/veterinary network between laboratories to enhance the listeriosis surveillance system in the Sicily region

An Italian Neurology Outpatient Clinic Facing SARS-CoV-2 Pandemic: Data From 2,167 Patients

Objective:Neurological sequelae of SARS-CoV-2 infection have already been reported, but there is insufficient data about the impact of the pandemic on the management of the patients with chronic neurological diseases. We aim to analyze the effect of COVID-19 pandemic and social restriction rules on these fragile patients. Methods:Patients with chronic neurologic diseases routinely followed at the outpatient clinic of Gemelli University Hospital, Rome, were assessed for symptoms suggestive of SARS-CoV-2 infection in the pandemic period, consequences of social restrictions, and neurological disease features, concomitant medical conditions, current medical and disease-specific treatments. Data source: a dedicated telephone survey designed to encompass questions on COVID-19 symptoms and on pandemic effects in chronic neurologic conditions. Results:Overall, 2,167 individuals were analyzed: 63 patients reported contact with COVID-19 positive cases, 41 performed the swab, and 2 symptomatic patients tested positive for COVID-19 (0.09%). One hundred fifty-eight individuals (7%) needed urgent neurological care, deferred due to the pandemic; 641 patients (30%) suspended hospital treatments, physiotherapy or other support interventions; 405 individuals (19%) reported a subjective worsening of neurological symptoms. Conclusions:In our population, the presence of neurological chronic diseases did not increase the prevalence of COVID-19 infection. Nevertheless, the burden of neurological disorders has been worsened by the lockdown