Acinetobacter baylyi Strain BD413 Can Acquire an Antibiotic Resistance Gene by Natural Transformation on Lettuce Phylloplane and Enter the Endosphere.

Antibiotic resistance spread must be considered in a holistic framework which comprises the agri-food ecosystems, where plants can be considered a bridge connecting water and soil habitats with the human microbiome. However, the study of horizontal gene transfer events within the plant microbiome is still overlooked. Here, the environmental strain Acinetobacter baylyi BD413 was used to study the acquisition of extracellular DNA (exDNA) carrying an antibiotic resistance gene (ARG) on lettuce phylloplane, performing experiments at conditions (i.e., plasmid quantities) mimicking those that can be found in a water reuse scenario. Moreover, we assessed how the presence of a surfactant, a co-formulant widely used in agriculture, affected exDNA entry in bacteria and plant tissues, besides the penetration and survival of bacteria into the leaf endosphere. Natural transformation frequency in planta was comparable to that occurring under optimal conditions (i.e., temperature, nutrient provision, and absence of microbial competitors), representing an entrance pathway of ARGs into an epiphytic bacterium able to penetrate the endosphere of a leafy vegetable. The presence of the surfactant determined a higher presence of culturable transformant cells in the leaf tissues but did not significantly increase exDNA entry in A. baylyi BD413 cells and lettuce leaves. More research on HGT (Horizontal Gene Transfer) mechanisms in planta should be performed to obtain experimental data on produce safety in terms of antibiotic resistance.

Zooplankton as a Transitional Host for Escherichia coli in Freshwater.

This study shows that Escherichia coli can be temporarily enriched in zooplankton under natural conditions and that these bacteria can belong to different phylogroups and sequence types (STs), including environmental, clinical, and animal isolates. We isolated 10 E. coli strains and sequenced the genomes of two of them. Phylogenetically, the two isolates were closer to strains isolated from poultry meat than to freshwater E. coli, albeit their genomes were smaller than those of the poultry isolates. After isolation and fluorescent protein tagging of strains ED1 and ED157, we show that Daphnia sp. can take up these strains and release them alive again, thus becoming a temporary host for E. coli. In a chemostat experiment, we show that this association does not prolong bacterial long-term survival, but at low abundances it also does not significantly reduce bacterial numbers. We demonstrate that E. coli does not belong to the core microbiota of Daphnia, suffers from competition by the natural Daphnia microbiota, but can profit from its carapax to survive in water. All in all, this study suggests that the association of E. coli with Daphnia is only temporary, but the cells are viable therein, and this might allow encounters with other bacteria for genetic exchange and potential genomic adaptation to the freshwater environment. IMPORTANCE The contamination of freshwater with feces-derived bacteria is a major concern regarding drinking water acquisition and recreational activities. Ecological interactions promoting their persistence are still very scarcely studied. This study, which analyses the survival of E. coli in the presence of zooplankton, is thus of ecological and water safety relevance.

Early Post-operative Feeding: An Investigation of Early Functional Outcomes for Oral Cancer Patients Treated with Surgical Resection and Free Flap Reconstruction.

Traditionally patients can remain nil by mouth (NBM) for up to 12 days after oral tumour resection with free flap reconstruction to reduce the risk of flap dehiscence, poor healing and fistulae. The literature reports that patients could on average remain an inpatient for up to 20 days post-surgery. An evaluation of the impact of a defined early oral feeding protocol was undertaken investigating functional outcomes and complications rates. We prospectively reviewed tracheostomy use, length of hospital stay, non-oral feeding status and swallowing function using the Performance Status Scale for Head and Neck Cancer (PSS-HN) within a defined early feeding protocol. Twenty-nine patients underwent surgical resection with free flap reconstruction for advanced primary oral cancer between January 2018 and December 2019. Average age was 59.5 (range 24-88). Tumour sites included oral tongue (n = 10), maxilla (n = 6), mandible (n = 6), floor of mouth (n = 5) and buccal mucosa (n = 2). Median time to decannulation was 7 days (range 3-20 days, n = 11). The majority of patients were able to tolerate at least oral fluids on day 1 post-operatively (86%, n = 25). In addition to oral intake, non-oral feeding was required in 90% (n = 26), the majority of which included a nasogastric tube (NGT) placed intraoperatively 54% (n = 14), others required gastrostomy 46% (n = 12). Median time to nasogastric tube removal was 6 days (range 3-15 days). Median length of hospital stay was 10 days (range 3-51). Mean PSS-Normalcy of Diet (NOD) score at point of hospital discharge was 36.55 (95% CI 30.9-42.2). Flap failure was noted in 3% (n = 1). The adoption of an early oral feeding protocol suggests that there is the potential for a shorter hospital stay and earlier swallowing rehabilitation.

Speech and Swallowing Outcomes Following Surgical Resection with Immediate Free Tissue Transfer Reconstruction for Advanced Osteoradionecrosis of the Mandible Following Radiation Treatment for Head and Neck Cancer.

Despite recent advances in the radiation techniques used for the treatment of head and neck cancer (HNC) including intensity-modulated radiotherapy (IMRT), mandibular osteoradionecrosis (ORN) remains a significant complication. Advanced stage ORN is managed surgically with resection and immediate free tissue transfer reconstruction. An evaluation of the functional speech and swallowing outcomes was undertaken for patients undergoing surgical management of advanced ORN. We retrospectively reviewed consecutive patients, at a single, tertiary cancer centre, who underwent surgical resection for advanced Notani grade III ORN. Outcomes investigated included use and duration of tracheostomy and swallowing and speech status using Performance Status Scale for Head and Neck Cancer Normalcy of Diet (PSS-NOD) and Understandability of Speech (PSS-Speech) at baseline and 3 months following surgery. Ten patients underwent surgical resection with free tissue transfer reconstruction between January 2014 and December 2019. Two patients required supplemental nutrition via a gastrostomy at three months post surgery. As per the PSS-NOD data half of the patients' (n = 5) diet remained stable (n = 2) or improved (n = 3) and half of the participants experienced a decline in diet (n = 5). The majority of patients had no speech difficulties at baseline (n = 8). The majority of patients' speech remained stable (n = 8) with two patients experiencing a deterioration in speech clarity following surgery. Well-designed studies with robust, sensitive multidimensional dysphagia and communication assessments are required to fully understand the impact of surgical management of advanced ORN using resection with free tissue transfer reconstruction.

A multi-residue analytical method for extraction and analysis of pharmaceuticals and other selected emerging contaminants in sewage sludge.

Sewage sludge is a by-product of wastewater treatment processes, and may be employed in agriculture as a fertilizer or in forestry for land reclamation. It is an important source of nutrients but its reuse can arouse concern on account of the wide range of contaminants that are retained and may persist during treatments. Information on the emerging contaminants (ECs) in sewage sludge in Italy is limited. The present study developed and applied a reliable analytical method for the analysis of 44 ECs in sewage sludge. ECs were extracted by accelerated solvent extraction followed by a clean-up step on solid-phase cartridges. High performance liquid chromatography coupled to mass spectrometry was used for analysis. The ECs, selected on the basis of their use and documented presence in the environment, were 42 pharmaceuticals belonging to 12 therapeutic categories and 2 perfluorinated substances. The method performance was good, with recoveries higher than 70%, good repeatability (<20%) and sensitivity in the low ng g-1 range, allowing measurement of the analytes selected. The method was applied for analysis of sludge from 12 wastewater treatment plants in Italy. The most abundant compounds were antibiotics, anti-inflammatories and antihypertensives and ranged up to 5 µg g-1 (ciprofloxacin). Seasonal differences were found for some antibiotics and anti-inflammatory drugs as well as some differences - in terms of concentration - with other European countries. This is the first Italian study to investigate the presence of a large number of ECs in sewage sludge and the results may be useful to drive future regulatory actions.

The impact of early oral feeding following head and neck free flap reconstruction on complications and length of stay.

BACKGROUND: Although the concept of maintaining a period of 'nil by mouth' following head and neck reconstruction is commonly held ideas on delaying function have changed dramatically since the introduction of peri-operative care recovery programmes. This study sought to evaluate the outcomes of early feeding on patients undergoing free flap reconstruction of oral defects with particular emphasis on post-operative complications and length of stay. METHODS: Data was gathered prospectively on two cohorts of patients treated in a tertiary referral centre comparing those undergoing a five-day post-operative period of 'nil by mouth' with a second group in which the aim was to start fluids and soft diet on the day following surgery. Complications and length of stay were evaluated. RESULTS: Both early and late feeding groups comprised of 200 patients. No significant differences were observed in terms of age, gender, smoking and alcohol use or tumour T and N stages between the two groups. 8% of patients had complications at the recipient site but no difference was observed in the rates of flap dehiscence or fistula formation between the two groups. Early feeding was associated with a statistically reduced length of hospital stay (mean 11.6 days vs 20.6 days, p < 0.01). CONCLUSIONS: Early oral feeding following head and neck free flap reconstruction is not associated with an increase incidence of peri-operative complications but reduces hospital stay. The latter may have far-reaching implications for patients' physical and psychological well-being in addition to health care resources.

An Environmental Escherichia coli Strain Is Naturally Competent to Acquire Exogenous DNA.

The diffusion of antibiotic resistance determinants in different environments, e.g., soil and water, has become a public concern for global health and food safety and many efforts are currently devoted to clarify this complex ecological and evolutionary issue. Horizontal gene transfer (HGT) has an important role in the spread of antibiotic resistance genes (ARGs). However, among the different HGT mechanisms, the capacity of environmental bacteria to acquire naked exogenous DNA by natural competence is still poorly investigated. This study aimed to characterize the ability of the environmental Escherichia coli strain ED1, isolated from the crustacean Daphnia sp., to acquire exogenous DNA by natural competence. Transformation experiments were carried out varying different parameters, i.e., cell growth phase, amount of exogenous DNA and exposition to artificial lake water (ALW) and treated wastewater to mimic environmental-like conditions that may be encountered in the agri-food system. Results were compared with those showed by the laboratory E. coli strain DH5α. Our experimental data, supported by genomic sequencing, showed that, when exposed to pure water, ED1 strain was able to acquire exogenous DNA with frequencies (10-8-10-9) statistically higher than the ones observed for DH5α strain (10-10). Interestingly, higher values were retrieved for ED1 than DH5α strains exposed to ALW (10-7 vs. 10-9, respectively) or treated wastewater (10-8 vs. 10-10, respectively). We tested, therefore, ED1 strain ability to colonize the rhizosphere of lettuce, a model plant representative of raw-consumed vegetables of high economic importance in the ready-to-eat food industry. Results showed that ED1 strain was able to efficiently colonize lettuce rhizosphere, revealing a stable colonization for 14 days-long period. In conclusion, ED1 strain ability to acquire exogenous DNA in environmental-like conditions by natural competence, combined with its ability to efficiently and stably colonize plant rhizosphere, poses the attention to food and human safety showing a possible route of diffusion of antibiotic resistance in the agri-food system, sustaining the "One Health" warnings related to the antibiotic spread.

Integrated surgery and radiology: trans-oral robotic surgery guided by real-time radiologist-operated intraoral ultrasound.

PURPOSE: We aimed to evaluate the feasibility and utility of intraoral ultrasound as a real-time guidance technique in trans-oral robotic surgery (TORS). METHODS: We report our early experiences of radiologist-operated intraoral ultrasound during TORS, providing information on tumour margin and important adjacent structures. Resection specimens were sonographically imaged for margin assessment. RESULTS: Four patients underwent ultrasound-guided TORS, with a close correlation between intraoperative and preoperative imaging in each case. The tumour was fully excised in three cases (one did not proceed due to difficult access). No ultrasound-related adverse events occurred, surgical margins were negative, and the treated patients are currently in a state of remission, with functional swallows. CONCLUSION: Intraoperative ultrasound can allow previously inaccessible disease to be robotically resected with confidence. Sonographic interrogation of the resected specimen correlated closely with histological margin analysis, and a novel technique of using saline to improve ultrasound conductance and therefore image quality is described.

Testing urban wastewater to assess compliance with prescription data through wastewater-based epidemiology: First case study in Italy.

Sub-optimal adherence to pharmacological therapy is one of the main reasons for poor effectiveness, reducing the patient's quality of life and affecting health-care economics. This study investigated the possibility for a wastewater-based epidemiology approach to assess the overall adherence of a population to some pharmacological therapies in a defined area. We selected specific active ingredients and their main urinary metabolites (biomarkers) according to the best practice protocol available and we measured them for the first time in urban wastewater. We conducted this first case study in Italy, considering the whole country and two Italian regions by sampling for five days six cities from north to south. Liquid chromatography tandem mass spectrometry was used to measure the biomarkers in raw wastewater, developing and validating specific analytical methods. We used concentrations measured in influent wastewater, together with a careful review of the excretion profile of the active ingredients selected, to back-calculate consumption (measured consumptions - MC) which were then compared with consumptions calculated from medical prescriptions (expected consumptions - EC) reported annually by the Italian Medicine Agency. In general there was a rough correlation between MC and EC, with ratios sometimes close to unity and always within a 0.3-3.0 range. Interpretation of the results suffers some biases, which are case-by-case discussed in detail, but inclusion of measurements of the urinary metabolites together with the active ingredients gave a fuller understanding of the results.

Pharmaceuticals and other contaminants in waters and sediments from Augusta Bay (southern Italy).

The contamination by pharmaceuticals products (PPs) in the marine environment is particularly relevant where wastewater treatment of urban areas on land is lacking. However, the number of studies focused on description of sources and fate of PP molecules in the marine environment remains still limited. In this study, the occurrence of 46 PPs was investigated in the marine and coastal-marine system (waters and sediments) of Augusta Bay (central Mediterranean Sea). This area is highly affected by industrial pollution and urban discharges (without wastewater treatment) and thus represents a 'natural laboratory' for exploring dynamics of multi-mixture contaminants in the marine environment. The study area is also part of the sub-region 'Central Mediterranean Sea' of the Marine Strategy Framework Directive and therefore offers an important reference site for exploring the distribution modes of PPs in the central Mediterranean Sea. In this work, samples of seawater, sediment, untreated wastewater, and marine receiving water were analysed using mass spectrometry with a target analysis for PPs and a suspect screening analysis for the presence of other contaminants. PPs concentration ranges were: 2426-67,155 ng/L for untreated wastewaters, 550-27,889 ng/L for marine receiving waters and 12-281 ng/L for seawaters. The highest concentrations were measured for the antibiotics, anti-inflammatories, cardiovascular and antihypertensive therapeutic classes. Likewise, sediments collected from untreated wastewater sewers resulted more contaminated. Ionic, non-ionic surfactants and personal care products were the most abundant compounds found in waters and sediments by suspect screening analysis. The risk associated with PPs contamination for aquatic organisms was relatively high in samples of marine receiving waters of the bay (with a risk quotient value up to 33,599). The levels of PPs in seawater and sediment compartments were generally not hazardous (RQ < 0.01), except for estrone with a calculated RQ = 2775.

Improved BSM sensitivity in diboson processes at linear colliders.

We study W + W - and Zh final sstates at future linear e + e - colliders; designing analyses specific to the various final state polarizations allows us to target specific beyond the Standard Model (BSM) effects, parametrized in the form of dimension-6 operators. We find that CLIC can access effects roughly an order of magnitude smaller than HL-LHC or ILC, and two orders of magnitude smaller than LEP. These results are interpreted in the context of well-motivated BSM scenarios-at weak and strong coupling-where we expect correlated effects in Drell-Yann processes. The latter turn out to have better discovery potential, although the diboson processes provide additional discriminating power, potentially furnishing a way to measure the spin and coupling of BSM states.

Transoral robotic surgery for recurrent cancers of the upper aerodigestive tract-Systematic review and meta-analysis.

BACKGROUND: Transoral robotic surgery (TORS) for recurrent head and neck (H&N) cancer is an emerging but relatively infrequent procedure. METHODS: Systematic review and meta-analysis of studies reporting survival data and functional outcomes for patients undergoing TORS for previously treated H&N cancers. RESULTS: Eight hundred seventy-eight records were identified, of which eight were eligible for inclusion, covering 161 cases (range 1-64). The pooled rates were as follows: 2-year overall survival 73.8% (4 studies, range 70.6-75.0, 95% confidence intervals (CI) 65.4 to 81.5, [I2 0.0%, P = 1.0]); 2-year disease-free survival 74.8% (4 studies, range 56.2-92.0, 95% CI 63.3 to 84.8, [I2 36.9%, P = .2]); postoperative hemorrhage 9.3% (4 studies, range 3.3-13.3, 95% CI 4.7 to 15.1, [I2 0.0%, P = .5]). CONCLUSIONS: Functional and oncological outcomes are favorable, although the follow-up is limited in the literature. Larger cohorts with longer follow-up are needed for definitive conclusions to be drawn.

Micropollutants in Lake Como water in the context of circular economy: A snapshot of water cycle contamination in a changing pollution scenario.

In this work we evaluated the contamination of the water cycle in Como Bay by measuring 38 selected pharmaceuticals in two main wastewater treatment plant in Switzerland and in Italy, two influents (River Breggia and Cosia), lake water (epilimnion and hypolimnion), as well as potable water. The collection of comparative information on the presence and environmental fate of these substances contributes to set specific environmental quality standard (EQS). The results presented show that the contamination of the lake reflects national health policies, which deeply influence the usage of chemicals. The outcomes of this study give an overall picture of contamination in the area, showing that concentrations of the measured compounds are generally low and under the commonly adopted ecotoxicological and toxicological thresholds. Only in a few cases did the contamination appear to be noteworthy, for some of the most persistent compounds (antibiotic macrolides, diclofenac, irbesartan, carbamazepine and dihydrocarbamazepine, bezafibrate, furosemide and hydrochlorothiazide). Some concern can be also be raised for the presence of antibiotics (clarithromycin) in drinking water, although at very low levels, due to the problem of antibiotic resistance.

Use of legal and illegal substances in Malé (Republic of Maldives) assessed by wastewater analysis.

This study used wastewater-based epidemiology (WBE) to investigate the lifestyle of the inhabitants of Malé, the capital of the Republic of Maldives. Raw wastewater 12-h composite samples were collected from nine pumping stations serving the city area - thus representative of the whole Malé population. Samples were analysed by liquid chromatography coupled to mass spectrometry for estimating the profile of use of a large number of substances including illicit drugs, alcohol, caffeine, tobacco and pharmaceuticals. The illicit drugs most used were cannabis (THC) and heroin (700 and 18 g/day), with lower consumption of cocaine and amphetamines (0.1-1.2 g/day). It is important to note that the consumption of cannabis in Malé was comparable to that measured in other countries, while the consumption of heroin was higher. Among cathinones, mephedrone was detected at the highest levels similar to other countries. Consumption of alcohol, which is not allowed in Maldives, was found (1.3 L/day/1000 inhabitants), but at a low level compared with other countries (6-44 L/day/1000 inhabitants), while the consumption of caffeine and tobacco was generally in line with reports from other countries. Unique information on pharmaceuticals use was also provided, since no official data were available. Human lifestyle was evaluated by applying for the first time the full set of WBE methodologies available in our laboratory. Results provided valuable epidemiological information, which may be useful for national and international agencies to understand population lifestyles better, including illicit drug issues, and for planning and evaluation of drug prevention programs in Malé.

Microbial assisted phytodepuration for water reclamation: Environmental benefits and threats.

Climate changes push for water reuse as a priority to counteract water scarcity and minimize water footprint especially in agriculture, one of the highest water consuming human activities. Phytodepuration is indicated as a promising technology for water reclamation, also in the light of its economic and ecological sustainability, and the use of specific bacterial inocula for microbial assisted phytodepuration has been proposed as a further advance for its implementation. Here we provided an overview on the selection and use of plant growth promoting bacteria in Constructed Wetland (CW) systems, showing their advantages in terms of plant growth support and pollutant degradation abilities. Moreover, CWs are also proposed for the removal of emerging organic pollutants like antibiotics from urban wastewaters. We focused on this issue, still debated in the literature, revealing the necessity to deepen the knowledge on the antibiotic resistance spread into the environment in relation to treated wastewater release and reuse. In addition, given the presence in the plant system of microhabitats (e.g. rhizosphere) that are hot spot for Horizontal Gene Transfer, we highlighted the importance of gene exchange to understand if these events can promote the diffusion of antibiotic resistance genes and antibiotic resistant bacteria, possibly entering in the food production chain when treated wastewater is used for irrigation. Ideally, this new knowledge will lead to improve the design of phytodepuration systems to maximize the quality and safety of the treated effluents in compliance with the 'One Health' concept.

Measuring Higgs Couplings without Higgs Bosons.

The measurement of Higgs couplings constitutes an important part of present standard model precision tests at colliders. We show that modifications of Higgs couplings induce energy-growing effects in specific amplitudes involving longitudinally polarized vector bosons, and we initiate a novel program to study these effects off shell and at high energy, rather than on the Higgs resonance. Our analysis suggests that these channels are complementary and competitive with familiar on-shell measurements; moreover, they offer endless opportunities for refinement and improvements.

Imaging Soft-tissue Sarcomas of the Head and Neck: A Tertiary Soft-tissue Sarcoma Unit Experience.

BACKGROUND/AIM: To describe imaging features of head and neck soft-tissue sarcomas. PATIENTS AND METHODS: Patients with a diagnosis of head and neck sarcoma between 2011 and 2015 were reviewed. RESULTS: There were a total of 62 patients (24 female; median age=60 years). Most common sarcomas were angiosarcoma, undifferentiated pleomorphic sarcoma and sarcoma not otherwise specified. They were most commonly located in cranial and neck superficial soft tissues. Average tumour size at presentation was 45 mm. One patient had metastasis at presentation (rhabdomyosarcoma); two had nodal disease (rhabdomyosarcoma and angiosarcoma) and two tumours contained calcification (chondrosarcoma and synovial sarcoma). Four arose after prior radiotherapy. CONCLUSION: Unlike the more common diagnosis of squamous cell carcinoma, the majority of head and neck sarcomas present as large, solitary, superficial masses without lymph node enlargement. Identification of these features on imaging should raise suspicion of a sarcoma diagnosis, particularly in the setting of previous irradiation or genetic susceptibility.

Risk assessment of a mixture of emerging contaminants in surface water in a highly urbanized area in Italy.

A complex mixture of emerging contaminants (ECs) occurs in the environment, with potential effects for aquatic organisms and human health. This study assessed the environmental risk of a mixture of ECs detected in the most urbanized and industrialized area of Italy. Water samples were collected in the Lambro river basin to the north and south of Milan, and were analyzed by liquid chromatography tandem mass spectrometry. The Environmental Risk Assessment (ERA) was performed calculating the Risk Quotients (RQs) for each EC as the ratio of Measured Concentrations (MECs) and Predicted No Effect Concentrations. The ERA was also conducted for the whole mixture of ECs by considering the RQs of the components. The results confirmed that ECs are ubiquitous in urbanized areas. The ERA for the single pollutants identified a panel of substances of environmental concern (estrogens, amoxicillin, clarythromycin, triclosan and nicotine). The ERA for the mixture indicated a potential cumulative risk for the substances that individually could be considered safe, highlighting the importance of taking the whole mixture of ECs into account for the ERA. This information may help establish EU regulations for ECs and environmental quality standards for regulatory purposes.

Data on occurrence and fate of emerging contaminants in a urbanised area.

These data and analyses support the research article "Mass balance of emerging contaminants in the water cycle of an highly urbanized and industrialized area of Italy" by Castiglioni et al. (2018) [1]. The occurrence of 80 emerging contaminats in waste and surface water was investigated in an highly urbanised area of Italy, the River Lambro basin. The data presented here include: (1) concentrations in untreated and treated wastewater of different wastewater treatment plants (WWTPs); (2) concentrations in surface water collected along the river Lambro, in the north and south of the city of Milan (main urban center in the area). These concentrations indicate the distribution and fate of emerging contaminats in the environment.

Beyond Positivity Bounds and the Fate of Massive Gravity.

We constrain effective field theories by going beyond the familiar positivity bounds that follow from unitarity, analyticity, and crossing symmetry of the scattering amplitudes. As interesting examples, we discuss the implications of the bounds for the Galileon and ghost-free massive gravity. The combination of our theoretical bounds with the experimental constraints on the graviton mass implies that the latter is either ruled out or unable to describe gravitational phenomena, let alone to consistently implement the Vainshtein mechanism, down to the relevant scales of fifth-force experiments, where general relativity has been successfully tested. We also show that the Galileon theory must contain symmetry-breaking terms that are at most one-loop suppressed compared to the symmetry-preserving ones. We comment as well on other interesting applications of our bounds.