Effective elimination of Staphylococcal contamination from hospital surfaces by a bacteriophage-probiotic sanitation strategy: a monocentric study.

Persistent contamination of hospital surfaces and antimicrobial resistance (AMR) are recognized major causes of healthcare-associated infections (HAI). We recently showed that a probiotic-based sanitation (PCHS) can stably decrease surface pathogens and reduce AMR and HAIs. However, PCHS action is slow and non-specific. By contrast, bacteriophages have been proposed as a decontamination method as they can rapidly attack specific targets, but their routine application has never been tested. Here we analyzed the feasibility and effectiveness of phage addition to PCHS sanitation, aiming to obtain a rapid and stable abatement of specific pathogens in the hospital environment. Staphylococcal contamination in the bathrooms of General Medicine wards was analyzed, being such areas the most contaminated and Staphylococci the most prevalent bacteria in such settings. Results showed that a daily phage application by nebulization induced a rapid and significant decrease of Staphylococcus spp. load on treated surfaces, up to 97% more than PCHS alone (p<0.001), suggesting that such system might be considered as a part of prevention and control strategies, to counteract outbreaks of specific pathogens and prevent associated infections

Pathogen Control in the Built Environment: A Probiotic-Based System as a Remedy for the Spread of Antibiotic Resistance

The high and sometimes inappropriate use of disinfectants and antibiotics has led to alarming levels of Antimicrobial Resistance (AMR) and to high water and hearth pollution, which today represent major threats for public health. Furthermore, the current SARS-CoV-2 pandemic has deeply influenced our sanitization habits, imposing the massive use of chemical disinfectants potentially exacerbating both concerns. Moreover, super-sanitation can profoundly influence the environmental microbiome, potentially resulting counterproductive when trying to stably eliminate pathogens. Instead, environmentally friendly procedures based on microbiome balance principles, similar to what applied to living organisms, may be more effective, and probiotic-based eco-friendly sanitation has been consistently reported to provide stable reduction of both pathogens and AMR in treated-environments, compared to chemical disinfectants. Here, we summarize the results of the studies performed in healthcare settings, suggesting that such an approach may be applied successfully also to non-healthcare environments, including the domestic ones, based on its effectiveness, safety, and negligible environmental impact

Introduction of NGS in Environmental Surveillance for Healthcare-Associated Infection Control

The hospital environment significantly contributes to the onset of healthcare associated infections (HAIs), representing the most frequent and severe complications related to health care. The monitoring of hospital surfaces is generally addressed by microbial cultural isolation, with some performance limitations. Hence there is need to implement environmental surveillance systems using more effective methods. This study aimed to evaluate next-generation sequencing (NGS) technologies for hospital environment microbiome characterization, in comparison with conventional and molecular methods, in an Italian pediatric hospital. Environmental samples included critical surfaces of randomized rooms, surgical rooms, intensive care units and delivery rooms. The resistome of the contaminating population was also evaluated. NGS, compared to other methods, detected with higher sensitivity the environmental bacteria, and was the only method able to detect even unsearched bacteria. By contrast, however, it did not detect mycetes, nor it could distinguish viable from dead bacteria. Microbiological and PCR methods could identify and quantify mycetes, in addition to bacteria, and PCR could define the population resistome. These data suggest that NGS could be an effective method for hospital environment monitoring, especially if flanked by PCR for species identification and resistome characterization, providing a potential tool for the control of HAI transmission

Evaluation of Anti-SARS-CoV-2 IgA Response in Tears of Vaccinated COVID-19 Subjects

Secretory IgA (sIgA), which may play an important role in the early defense against SARS-CoV-2 infection, were detected in the eye of COVID-19 patients. However, an evaluation of the sIgA response in the tears of vaccinated or non-vaccinated COVID-19 subjects is still lacking. Aimed at characterizing sIgA mucosal immunity in the eye, this study analyzed tear samples from 77 COVID-19 patients, including 63 vaccinated and 14 non-vaccinated subjects. The groups showed similar epidemiological features, but as expected, differences were observed in the percentage of asymptomatic/pauci-symptomatic subjects in the vaccinated vs. non-vaccinated cohort (46% and 29% of the total, respectively). Consistent with this, ocular sIgA values, evaluated by a specific quantitative ELISA assay, were remarkably different in vaccinated vs. non-vaccinated group for both frequency (69.8% vs. 57.1%, respectively) and titer (1372.3 U/mL vs. 143.7 U/mL, respectively; p = 0.01), which was significantly differently elevated depending on the type of administered vaccine. The data show for the first time significant differences of available vaccines to elicit sIgA response in the eye and suggest that quantitative tear-based sIgA tests may potentially serve as a rapid and easily accessible biomarker for the assessment of the development of a protective mucosal immunity toward SARS-CoV-2

Characterization of the Pathogenic Potential of the Beach Sand Microbiome and Assessment of Quicklime as a Remediation Tool

Beach sand may act as a reservoir for potential human pathogens, posing a public health risk. Despite this, the microbiological monitoring of sand microbiome is rarely performed to determine beach quality. In this study, the sand microbial population of a Northern Adriatic Sea beach sand was profiled by microbiological (CFU counts) and molecular methods (WGS, microarray), showing significant presence of potential human pathogens including drug-resistant strains. Consistent with these results, the potential of quicklime as a restoring method was tested in vitro and on-field. Collected data showed that adding 1-3% quicklime (w/w) to sand provided an up to -99% of bacteria, fungi, and viruses, in a dose- and time-dependent manner, till 45 days post-treatment. In conclusion, data suggest that accurate monitoring of sand microbiome may be essential, besides water, to assess beach quality and safety. Moreover, first evidences of quicklime potential for sand decontamination are provided, suggesting its usage as a possible way to restore the microbiological quality of sand in highly contaminated areas

Green procedure for one-pot synthesis of azelaic acid derivatives using metal catalysis

Background & Objective: A green one-pot synthesis of oleic acid (1) derivatives is promoted by Rare Earth Metal (REM) triflates and commercial Molybdenum dioxo dichloride (MoCl 2 O 2 ) in the presence hydrogen peroxide as a green oxidant. Results: The protocol permits to govern the oxidation selectivity by simply choosing the proper combination of Mo and Sc catalysts. Conclusion: Methyl oleate epoxide 2a and azelaic acid 6 thus obtained are valuable industrial intermediates for synthesizing bio-compostable plastics, plasticizers of PVC, lubricating oils, cosmetics and pharmaceuticals (bactericides, anti-inflammatories, etc.)

A highly integrated bionic hand with neural control and feedback for use in daily life

Restoration of sensorimotor function after amputation has remained challenging because of the lack of human-machine interfaces that provide reliable control, feedback, and attachment. Here, we present the clinical implementation of a transradial neuromusculoskeletal prosthesis-a bionic hand connected directly to the user's nervous and skeletal systems. In one person with unilateral below-elbow amputation, titanium implants were placed intramedullary in the radius and ulna bones, and electromuscular constructs were created surgically by transferring the severed nerves to free muscle grafts. The native muscles, free muscle grafts, and ulnar nerve were implanted with electrodes. Percutaneous extensions from the titanium implants provided direct skeletal attachment and bidirectional communication between the implanted electrodes and a prosthetic hand. Operation of the bionic hand in daily life resulted in improved prosthetic function, reduced postamputation, and increased quality of life. Sensations elicited via direct neural stimulation were consistently perceived on the phantom hand throughout the study. To date, the patient continues using the prosthesis in daily life. The functionality of conventional artificial limbs is hindered by discomfort and limited and unreliable control. Neuromusculoskeletal interfaces can overcome these hurdles and provide the means for the everyday use of a prosthesis with reliable neural control fixated into the skeleton

USE OF PROBIOTICS AND BACTERIOPHAGES IN HOSPITAL ENVIRONMENT FOR HAI CONTROL: POTENTIAL AS SANITIZING AGENTS

Use of probiotics and bacteriophages in hospital environment for HAI control: Potential as sanitizing agents. Healthcare Associated Infections (HAIs) are a global concern, further threatened by the increasing of multidrug resistant HAI-associated pathogens. Persistent contamination of hospital surfaces contributes to HAI transmission, and it is not efficiently controlled by conventional chemical-based cleaning. In the search for effective approaches, we recently studied an eco-sustainable probiotic-based cleaning system (Probiotic Cleaning Hygiene System, PCHS), showing that it can stably abate surface pathogens, without selecting antibiotic-resistant species. Based on such observations, the first aim of the project was to determine whether PCHS application could impact on HAI incidence. To this purpose, a multicentre, pre-post interventional study was performed for 18 months in the Internal Medicine wards of six Italian public hospitals. The intervention consisted uniquely in the substitution of conventional sanitation with PCHS, maintaining unaltered any other procedure influencing HAI control. For the whole duration of the study, HAI incidence and surface bioburden were analysed in parallel. PCHS was shown to be associated with a stable decrease of surface pathogens ( >80% more compared to pre-PCHS phase), accompanied by a concurrent drop of microbiota drug-resistance genes, confirming in a large sample the PCHS ability to effectively remodulate the environmental contamination and reduce the antimicrobial resistance (AMR) burden. Notably, the analysis of the HAI incidence evidenced that the PCHS-induced remodulation of hospital microbiota was associated with a 52% decrease of HAI incidence in PCHS compared to pre-PCHS period (P<0.0001). However, PCHS action was gradual and non-specific, and thus it cannot be considered as a rapid mean for the eradication of specifically targeted pathogens. With the aim of improving PCHS performance, we focussed upon the possibility to use the bacteriophage, which due to the selective killing of specific bacteria, have been proposed as a decontamination method in literature. To this aim, phage activity was first assessed in vitro and in situ, on different types of hard surfaces artificially contaminated by the major HAI-associated bacteria. The results showed that phages could rapidly and significantly reduce target bacteria on treated surfaces, and maintained their full activity when suspended in the detergent. With the aim to analyse the feasibility and effectiveness of a combined phage/PCHS sanitation in hospital, a monocentric study was performed in the Internal Medicine ward of an Italian hospital, focusing on the Staphylococcal contamination in the bathrooms, being bathrooms the most contaminated areas and Staphylococci the most prevalent bacteria in such settings. Eight total rooms were enrolled, four of which were treated with PCHS alone, whereas the other four rooms received combined phage/PCHS sanitation. Staphylococcal contamination was characterized prior phage application, to check the susceptibility of the contaminating strains, showing that all most prevalent species (coagulase–negative Staphylococci) were susceptible to phage lysis. Treated surfaces were analysed for residual Staphylococcal contamination and for phage presence by specific microbiological and molecular methods. Nebulization of phages in addition to PCHS sanitation was associated with a rapid, significant and specific decrease of Staphylococcus spp. on treated surfaces, up to 97% more than PCHS alone (P<0.001). No variations were observed in other bacterial genera, confirming a specific action of phages. Collected data suggest that the use of a phage/probiotic sanitation is associated with a rapid, specific and stable decontamination and such system might be considered as a part of infection prevention and control strategies, to counteract outbreaks of specific pathogens and prevent associated infections.Le infezioni correlate all'assistenza sanitaria (ICA) e l’aumento dei microrganismi multiresistenti ai farmaci rappresentano un problema globale. D’altra parte, la contaminazione persistente delle superfici ospedaliere contribuisce alla trasmissione dell'ICA, e non è controllata efficientemente dalla pulizia convenzionale. Nella ricerca di approcci ecosostenibili e maggiormente efficaci, abbiamo recentemente dimostrato che un sistema di pulizia a base probiotica (Probiotic Cleaning Hygiene System, PCHS) può abbattere stabilmente i patogeni presenti sulle superfici ospedaliere, senza selezionare specie antibiotico-resistenti. Sulla base dei risultati precedenti, il primo obiettivo è stato quello di determinare se l'applicazione del PCHS potesse avere un impatto sull'incidenza dell'ICA. A questo scopo, è stato eseguito uno studio multicentrico, pre-post interventistico, nei reparti di Medicina Interna di sei ospedali italiani per 18 mesi. L'intervento è consistito esclusivamente nella sostituzione della sanificazione chimica con il PCHS, mantenendo inalterata ogni altra procedura che potesse influire sul controllo delle ICA. Per tutta la durata dello studio, l'incidenza delle ICA e la contaminazione superficiale sono state analizzate in parallelo. Il PCHS è risultato associato ad una maggior e stabile diminuzione dei patogeni superficiali rispetto alla sanificazione convenzionale (diminuzione media dell'83%), accompagnata da un decremento di tutti i geni di resistenza ai farmaci presenti nel microbiota residuo, confermando anche in campioni più ampi l’effettiva rimodulazione del microbiota da parte del sistema probiotico. Il PCHS è risultato associato ad una significativa diminuzione dell'incidenza di ICA (diminuzione del 52%, p<0,001) negli ospedali trattati, mostrando un ruolo protettivo. Tuttavia, l'azione del PCHS è graduale e non specifica. Per migliorare le prestazioni del sistema si è pensato all’uso di batteriofagi, che, essendo specifici e ad azione rapida, sono stati proposti in letteratura come metodo di decontaminazione. A questo scopo, l'attività dei fagi è stata prima valutata in vitro su diversi tipi di superfici dure artificialmente contaminate dai principali batteri ICA-associati. I risultati hanno mostrato che i fagi potevano ridurre rapidamente i batteri target sulle superfici trattate e mantenevano la loro piena attività quando sospesi nel detergente. Con l'obiettivo di analizzare l'efficacia di un trattamento combinato fagi/PCHS sul campo, è stato condotto uno studio monocentrico in un ospedale italiano, analizzando la contaminazione stafilococcica nei bagni, essendo questi ultimi le aree più contaminate e gli stafilococchi i batteri più prevalenti in tali ambienti. Sono state incluse otto camere in totale, quattro delle quali sono state trattate con solo il PCHS, mentre le altre con i fagi in aggiunta al PCHS. Prima di effettuare il trattamento, abbiamo valutato la sensibilità ai fagi delle specie contaminanti le superfici, e i risultati hanno mostrato che le specie più prevalenti (stafilococchi coagulasi negativi) erano sensibili all’azione litica virale. Durante lo studio, le superfici trattate sono state analizzate con metodi microbiologici e molecolari specifici. I risultati hanno evidenziato una rapida, e specifica diminuzione di stafilococchi sulle superfici trattate quando usato il sistema combinato (fino al 97% in più rispetto al trattamento con il PCHS (P<0,001)). Non sono state osservate variazioni in altri batteri, a conferma di un'azione specifica dei fagi. I dati suggeriscono che un sistema di sanificazione con fagi/PCHS determina una decontaminazione rapida, specifica e stabile, e potrebbe quindi essere considerato come parte delle strategie di prevenzione e controllo delle infezioni, per contrastare gli outbreaks causati da specifici agenti patogeni

Efficient removal of hospital pathogens from hard surfaces by a combined use of bacteriophages and probiotics: potential as sanitizing agents

Purpose Many hospital-acquired infections (HAI) can be transmitted by pathogens contaminating hospital surfaces, not efficiently controlled by conventional sanitation, which can indeed contribute to the selection of multidrug-resistant (MDR) strains. Bacteriophages have been suggested as decontaminating agents, based on their selective ability to kill specific bacteria. However, there are no data on their stability in detergents and their potential use in routine sanitation. On the other hand, a probiotic-based sanitation (Probiotic Cleaning Hygiene System, PCHS) was recently shown to stably reduce pathogens on treated surfaces. However, its action is not specific and slow, being based on competitive antagonism. This work was aimed to assess the effectiveness of a combined use of phages and PCHS in removing HAI-associated pathogens from different hard surfaces. Methods The decontamination ability of phages in PCHS was tested in vitro and in situ, against drug-susceptible or resistant S. aureus, E. coli and P. aeruginosa strains, and using bacterial densities similar to those detected on hospital surfaces. Results Phages targeted efficiently all tested bacteria, maintaining their full activity when added to PCHS detergent. Notably, the combined use of phages and PCHS not only resulted in a rapid reduction (up to >90%) of the targeted pathogens, but, due to the stabilizing effect of probiotics, the pathogens were maintained at low levels (>99%) also at later times, when instead the effect of phages tends to diminish. Conclusion These results suggest that a combined biological system might be successfully used in hospital sanitation protocols, potentially leading to an effective and safe elimination of MDR pathogens from the hospital environment

HHV-6A/6B infection of NK cells modulates the expression of miRNAs and transcription factors potentially associated to impaired NK activity

Natural killer (NK) cells have a critical role in controlling virus infections, and viruses have evolved several mechanisms to escape NK cell functions. In particular, Human herpesvirus 6 (HHV-6) is associated with diseases characterized by immune dysregulation and has been reported to infect NK cells. We recently found that HHV-6 in vitro infection of human thyroid follicular epithelial cells and T-lymphocytes modulates several miRNAs associated with alterations in immune response. Since miRNAs are key regulators of many immune pathways, including NK cell functions, we aimed to study the impact of HHV-6A and -6B in vitro infection on the intracellular mediators correlated to NK cell function. To this purpose, a human NK cell line (NK-92) was infected in vitro with HHV-6A or 6B and analyzed for alterations in the expression of miRNAs and transcription factors. The results showed that both viruses establish lytic replication in NK-92 cells, as shown by the presence of viral DNA, expression of lytic transcripts and antigens, and by the induction of an evident cytopathic effect. Notably, both viruses, although with species-specific differences, induced significant modifications in miRNA expression of miRNAs known for their role in NK cell development, maturation and effector functions (miR-146, miR-155, miR-181, miR-223), and on at least 13 miRNAs with recognized role in inflammation and autoimmunity. Also the expression of transcription factors was significantly modified by HHV-6A/6B infection, with an early increase of ATF3, JUN and FOXA2 by both species, whereas HHV-6A specifically induced a 15-fold decrease of POU2AF1, and HHV-6B an increase of FOXO1 and a decrease of ESR1. Overall, our data show that HHV-6A and -6B infections have a remarkable effect on the expression of miRNAs and transcription factors, which might be important in the induction of NK cell function impairment, virus escape strategies and related pathologies