Comparing Visual and Software-Based Quantitative Assessment Scores of Lungs' Parenchymal Involvement Quantification in COVID-19 Patients.

(1) Background: Computed tomography (CT) plays a paramount role in the characterization and follow-up of COVID-19. Several score systems have been implemented to properly assess the lung parenchyma involved in patients suffering from SARS-CoV-2 infection, such as the visual quantitative assessment score (VQAS) and software-based quantitative assessment score (SBQAS) to help in managing patients with SARS-CoV-2 infection. This study aims to investigate and compare the diagnostic accuracy of the VQAS and SBQAS with two different types of software based on artificial intelligence (AI) in patients affected by SARS-CoV-2. (2) Methods: This is a retrospective study; a total of 90 patients were enrolled with the following criteria: patients' age more than 18 years old, positive test for COVID-19 and unenhanced chest CT scan obtained between March and June 2021. The VQAS was independently assessed, and the SBQAS was performed with two different artificial intelligence-driven software programs (Icolung and CT-COPD). The Intraclass Correlation Coefficient (ICC) statistical index and Bland-Altman Plot were employed. (3) Results: The agreement scores between radiologists (R1 and R2) for the VQAS of the lung parenchyma involved in the CT images were good (ICC = 0.871). The agreement score between the two software types for the SBQAS was moderate (ICC = 0.584). The accordance between Icolung and the median of the visual evaluations (Median R1-R2) was good (ICC = 0.885). The correspondence between CT-COPD and the median of the VQAS (Median R1-R2) was moderate (ICC = 0.622). (4) Conclusions: This study showed moderate and good agreement upon the VQAS and the SBQAS; enhancing this approach as a valuable tool to manage COVID-19 patients and the combination of AI tools with physician expertise can lead to the most accurate diagnosis and treatment plans for patients.

Orange Peel Waste as Feedstock for the Production of Glycerol-Free Biodiesel by the Microalgae Nannochloropsis oculata.

The bioconversion of agri-food waste into high-value products is gaining growing interest worldwide. Orange peel waste (OPW) is the main by-product of orange juice production and contains high levels of moisture and carbohydrates. In this study, the orange waste extract (OWE) obtained through acid hydrolysis of OPW was used as a substrate in the cultivation of the marine microalgae Nannochloropsis oculata. Photoheterotrophic (PH) and Photoautotrophic (PA) cultivations were performed in OWE medium and f/2 medium (obtained by supplementing OWE with macro- and micronutrients of f/2 medium), respectively, for 14 days. The biomass yields in PA and PH cultures were 390 mg L-1 and 450 mg L-1, while oil yields were 15% and 28%, respectively. The fatty acid (FA) profiles of PA cultures were mostly represented by saturated (43%) and monounsaturated (46%) FAs, whereas polyunsaturated FAs accounted for about 10% of the FAs. In PH cultures, FA profiles changed remarkably, with a strong increase in monounsaturated FAs (77.49%) and reduced levels of saturated (19.79%) and polyunsaturated (2.72%) FAs. Lipids obtained from PH cultures were simultaneously extracted and converted into glycerol-free biodiesel using an innovative microwave-assisted one-pot tandem protocol. FA methyl esters were then analyzed, and the absence of glycerol was confirmed. The FA profile was highly suitable for biodiesel production and the microwave-assisted one-pot tandem protocol was more effective than traditional extraction techniques. In conclusion, N. oculata used OWE photoheterotrophically, resulting in increased biomass and oil yield. Additionally, a more efficient procedure for simultaneous oil extraction and conversion into glycerol-free biodiesel is proposed.

How healthcare workers reacted to the different COVID-19 waves: An Italian survey.

INTRODUCTION: The COVID-19 pandemic had a huge impact on radiology departments all over the world, affecting both management and healthcare workers (HCWs). Therefore, it became challenging to guarantee high standards of diagnosis while keeping up with the workload. METHODS: The study was approved by the institutional review board. Its aim was to assess the impact of the COVID-19 pandemic over the radiology departments and HCWs through a survey. The questionnaire was available online from January to March 2022. Twelve areas of interest (sessions) were highlighted in the survey. RESULTS: The number of total responders was 1376 and 73.7% of participants worked in public healthcare facilities. Comparisons between participants working in public versus private healthcare facilities were carried out using chi-square tests and Fisher tests. Within public healthcare workers, 82% affirmed having operating instruction protocols regarding confirmed or suspected COVID-19 patient CT management (p< 0.001). Private healthcare facilities had fewer CT scanners available in general (p< 0.001); in fact, only 18% of them affirmed having two or more CT scanners, and did not have CT scanners dedicated to confirmed or suspected COVID-19 patients (p< 0.001). Finally, public facilities strongly reduced (by 88%) the number of examinations booked during the first wave, compared to private healthcare facilities (p< 0.001). CONCLUSION: This survey showed that public facilities appeared to be better prepared from an organizational point of view than private facilities. Rescheduling the examinations booked during the first COVID-19 wave was challenging and not always possible.

Anti-Bacterial Adhesion on Abiotic and Biotic Surfaces of the Exopolysaccharide from the Marine Bacillus licheniformis B3-15.

The eradication of bacterial biofilm represents a crucial strategy to prevent a clinical problem associated with microbial persistent infection. In this study we evaluated the ability of the exopolysaccharide (EPS) B3-15, produced by the marine Bacillus licheniformis B3-15, to prevent the adhesion and biofilm formation of Pseudomonas aeruginosa ATCC 27853 and Staphylococcus aureus ATCC 29213 on polystyrene and polyvinyl chloride surfaces. The EPS was added at different times (0, 2, 4 and 8 h), corresponding to the initial, reversible and irreversible attachment, and after the biofilm development (24 or 48 h). The EPS (300 µg/mL) impaired the initial phase, preventing bacterial adhesion even when added after 2 h of incubation, but had no effects on mature biofilms. Without exerting any antibiotic activity, the antibiofilm mechanisms of the EPS were related to the modification of the (i) abiotic surface properties, (ii) cell-surface charges and hydrophobicity, and iii) cell-to-cell aggregation. The addition of EPS downregulated the expression of genes (lecA and pslA of P. aeruginosa and clfA of S. aureus) involved in the bacterial adhesion. Moreover, the EPS reduced the adhesion of P. aeruginosa (five logs-scale) and S. aureus (one log) on human nasal epithelial cells. The EPS could represent a promising tool for the prevention of biofilm-related infections.

A Novel Serum-Based Diagnosis of Alzheimer's Disease Using an Advanced Phage-Based Biochip.

55 million people worldwide suffer from Alzheimer's disease (AD). A definitive diagnosis of AD is made postmortem after a neuropathological examination of the brain. There is an urgent need for an innovative, noninvasive methodology that allows for an early and reliable diagnosis. Several engineered phages that recognized Aß-autoantibodies present in the sera of AD patients are previously identified. Here, novel phages are tested for their ability to accurately discriminate AD sera using immunophage-polymerase chain reaction in a miniatured biochip. It is found that five of the six phages analyzed discriminate between healthy controls and AD patients. Further, by combining the response of two phages, non-AD and severe AD cases are identified with 100% accuracy and mild-to-moderate cases with 90% accuracy. While the number of cases used here are relatively small and can be confirmed in larger cohorts, this first-of-a-kind system represents an innovative methodology with the potential of having a major impact in the AD field: from a clinical perspective, it can aid physicians in making an accurate AD diagnosis; from a research perspective, it can be used as a surrogate for AD clinical trials.

Improvement of Biomass and Phycoerythrin Production by a Strain of Rhodomonas sp. Isolated from the Tunisian Coast of Sidi Mansour.

Microalgae are photoautotrophic microorganisms known as producers of a large variety of metabolites. The taxonomic diversity of these microorganisms has been poorly explored. In this study, a newly isolated strain was identified based on the 18S rRNA encoding gene. The phylogenetic analysis showed that the isolated strain was affiliated with the Rhodomonas genus. This genus has greatly attracted scientific attention according to its capacity to produce a large variety of metabolites, including phycoerythrin. Growth and phycoerythrin production conditions were optimized using a Plackett-Burman design and response surface methodology. An expression profile analysis of the cpeB gene, encoding the beta subunit of phycoerythrin, was performed by qRT-PCR under standard and optimized culture conditions. The optimization process showed that maximum cell abundance was achieved under the following conditions: CaCl2 = 2.1328 g/L, metal solution = 1 mL/L, pH = 7 and light intensity = 145 µmol photons/m2/s, whereas maximum phycoerythrin production level occurred when CaCl2 = 1.8467 g/L, metal solution = 1 mL/L, pH = 7 and light intensity = 157 µmol/m2/s. In agreement, positive transcriptional regulation of the cpeB gene was demonstrated using qRT-PCR. This study showed the successful optimization of abiotic conditions for highest growth and phycoerythrin production, making Rhodomonas sp. suitable for several biotechnological applications.

Different Lung Parenchyma Quantification Using Dissimilar Segmentation Software: A Multi-Center Study for COVID-19 Patients.

BACKGROUND: Chest Computed Tomography (CT) imaging has played a central role in the diagnosis of interstitial pneumonia in patients affected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and can be used to obtain the extent of lung involvement in COVID-19 pneumonia patients either qualitatively, via visual inspection, or quantitatively, via AI-based software. This study aims to compare the qualitative/quantitative pathological lung extension data on COVID-19 patients. Secondly, the quantitative data obtained were compared to verify their concordance since they were derived from three different lung segmentation software. METHODS: This double-center study includes a total of 120 COVID-19 patients (60 from each center) with positive reverse-transcription polymerase chain reaction (RT-PCR) who underwent a chest CT scan from November 2020 to February 2021. CT scans were analyzed retrospectively and independently in each center. Specifically, CT images were examined manually by two different and experienced radiologists for each center, providing the qualitative extent score of lung involvement, whereas the quantitative analysis was performed by one trained radiographer for each center using three different software: 3DSlicer, CT Lung Density Analysis, and CT Pulmo 3D. RESULTS: The agreement between radiologists for visual estimation of pneumonia at CT can be defined as good (ICC 0.79, 95% CI 0.73-0.84). The statistical tests show that 3DSlicer overestimates the measures assessed; however, ICC index returns a value of 0.92 (CI 0.90-0.94), indicating excellent reliability within the three software employed. ICC was also performed between each single software and the median of the visual score provided by the radiologists. This statistical analysis underlines that the best agreement is between 3D Slicer "LungCTAnalyzer" and the median of the visual score (0.75 with a CI 0.67-82 and with a median value of 22% of disease extension for the software and 25% for the visual values). CONCLUSIONS: This study provides for the first time a direct comparison between the actual gold standard, which is represented by the qualitative information described by radiologists, and novel quantitative AI-based techniques, here represented by three different commonly used lung segmentation software, underlying the importance of these specific values that in the future could be implemented as consistent prognostic and clinical course parameters.

COVID-19 outbreak impact on health professionals: A survey on the Italian radiographer experience.

AIM: To evaluate the impact of the Phase 1 COVID-19 (C19) outbreak on Italian Radiographers. MATERIAL AND METHODS: COVID-19 has spread rapidly worldwide. Many patients underwent radiological examinations, leading to a high risk of infection within the radiology department's staff. Italy was the first-hit European country to face the COVID-19 outbreak and the impact on radiographers was huge. An online survey was disseminated to investigate the involvement and working environment of Italian radiographers during the first outbreak of COVID-19. RESULTS: Of the 840 responders, 65% were men. The majority of the responding Health-care Workers (HCW) was represented by radiographers (96%), from high-prevalence regions (82%; p<.05). Forty-five percent were involved in the activation of the protocol for the management of COVID-19 positive patients, without exhaustive indication for Plain Radiography and Computed Tomography (CT). Only 17% of hospitals counted on available guidelines for serious infections (p<0.05). Diagnostic examinations were mainly performed by a single radiographer (62%). Many professionals (69%) confirmed wearing all indispensable PPE in case of COVID-19 positive patients. CONCLUSION: The primary objective of management strategies should be to redact standardized policies for the safeguarding of patient's health and operator's safety. All front-line workers, including radiographers working in diagnostic services, should be involved in the decision-making process to generate wellness and awareness.

Evaluation of Betacoronavirus OC43 and SARS-CoV-2 Elimination by Zefero Air Sanitizer Device in a Novel Laboratory Recirculation System.

Indoor air sanitizers contrast airborne diseases and particularly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)/Coronavirus disease 2019 (COVID-19). The commercial air sanitizer Zefero (Cf7 S.r.l., San Giovanni La Punta, Italy) works alternatively using a set of integrated disinfecting technologies (namely Photocatalysis/UV mode) or by generating ozone (Ozone mode). Here we evaluated the virucidal efficacy of Zefero setup modes against human Betacoronavirus OC43 and SARS-CoV-2. For this purpose, we designed a laboratory test system in which each virus, as aerosol, was treated with Photocatalysis/UV or Ozone mode and returned into a recirculation plexiglass chamber. Aerosol samples were collected after different times of exposure, corresponding to different volumes of air treated. The viral RNA concentration was determined by qRT-PCR. In Photocatalysis/UV mode, viral RNA of OC43 or SARS-CoV-2 was not detected after 120 or 90 min treatment, respectively, whereas in Ozone mode, viruses were eliminated after 30 or 45 min, respectively. Our results indicated that the integrated technologies used in the air sanitizer Zefero are effective in eliminating both viruses. As a reliable experimental system, the recirculation chamber developed in this study represents a suitable apparatus for effectively comparing the disinfection capacity of different air sanitizers.

Nutritional conditions of the novel freshwater Coccomyxa AP01 for versatile fatty acids composition.

AIMS: This study was to analyse the biomass production and fatty acids (FAs) profiles in a newly isolated chlorophyte, namely Coccomyxa AP01, under nutritionally balanced (NB) conditions (comparing nitrate and urea as nitrogen sources) and nitrogen or phosphate deprivation. METHODS AND RESULTS: Lipid yields was about 30%-40% of dried biomasses in all examined nutritional conditions. Under NB conditions, lipids were principally constituted by monounsaturated FAs, mainly represented by oleic acid, and saturated and polyunsaturated FAs at similar concentrations. Nutrients deprivation induced remarkable changes in FAs profiles, with the highest amounts of saturated (42%-46%), followed by similar amounts of monounsaturated and polyunsaturated, and the emergence of rare long-chain FAs. Under phosphate deprivation, biomass yield was similar to NB conditions, with the highest yield of saturated (mainly palmitic acid) and of polyunsaturated FAs (33%) (mainly linoleic and linolenic acids). CONCLUSIONS: Balanced or deprived nutritional conditions in Coccomyxa AP01 induced a selective production and composition of FAs. The phosphate-deprivation condition concomitantly provided high biomass yield and the production of high value saturated and polyunsaturated FAs with industrial interest. SIGNIFICANCE AND IMPACT OF THE STUDY: Coccomyxa AP01 could be considered a promising source of different FAs, including also docosapentaenoic acid, for several commercial purposes spanning from biodiesel production, pharmaceutical and cosmetic applications to innovative aquaculture fish feeds.

Glutamine-induced filamentous cells of Pseudomonas mediterranea CFBP-5447T as producers of PHAs.

Polyhydroxyalkanoates (PHAs) are considerable biopolymers that have gained an increasing biotechnological interest in different applications, although their industrial production presents several limitations. Filamentous bacterial cells could represent a possible strategy to increase PHA yield, since more abundant PHA inclusions can be stored in elongated than in rod-shaped cells. At first, we determined the optimal batch culture conditions to induce filamentation in Pseudomonas mediterranea CFBP-5447T, using glutamine, glycerol, glucose, and sodium octanoate, as the sole carbon source, at low- (100 rpm) or high- (250 rpm) shaking speeds. Successively, a fermentative process was set up using glutamine in a co-metabolic strategy with glycerol, and the PHAs production was compared in rod-shaped and filamentous cells. High glutamine concentrations (from 28 to 56 mM) were able to induce alone filamentation, whereas at lower glutamine concentrations (5-10 mM), the shaking speeds became critical to allow or not filamentous phenotype. PHA granule production was higher in filamentous than in rod-shaped cells, when glycerol (46.6 mM) was added to glutamine (5 mM) in co-metabolism, and fermentation was performed at a low-shaking speed. After extraction and precipitation, PHA yield was about two times higher in filamentous than that rod-shaped cells. Our results provide new insights into filament-inducing conditions and indicate a potential use of filamentous P. mediterranea CFBP-5447T cells to increase PHA yield. These findings could have great advantages in PHAs recovering during downstream processes, since the harvesting of elongated cells is much less time-consuming and energy expensive than required with rod-shaped cells.

Biosynthesis and structural characterization of polyhydroxyalkanoates produced by Pseudomonas aeruginosa ATCC 27853 from long odd-chain fatty acids.

Pseudomonas aeruginosa ATCC 27853 was cultured on media containing long odd-chain fatty acids. Heptadecanoic, nonadecanoic, and heneicosanoic acids sustained cell growth and resulted in polyhydroxyalkanoate (PHA) accumulation when culturing was conducted under nitrogen starvation conditions. No PHA was produced using a complete or magnesium-deprived medium. The isolated polyesters were characterized by gas chromatography and liquid chromatography-electrospray ionization mass spectrometry (ESI-MS) of methanolyzed samples, 1H and 13C NMR spectroscopy, gel permeation chromatography, ESI MS of partially pyrolyzed samples, and differential scanning calorimetry. These PHAs are composed of seven different odd-chain repeating units starting from 3-hydroxyvalerate, with the highest species being the, to date, unreported constituent 3-hydroxyheptadecanoate, and minor amounts of 2 or 3 even-chain comonomers. The PHAs are soft, sticky, rubber-like materials having glass transition temperatures between -45 and -39°C, melting temperatures between 48 and 52°C, enthalpies of melting around 11J/g, and molar masses ranging from 77 to 188kg/mol. Statistical analysis of the ESI mass spectra of the products of their partial pyrolysis showed that they are pure copolymers and not a blend of copolymers or homopolymers.

Carbon source effects on the mono/dirhamnolipid ratio produced by Pseudomonas aeruginosa L05, a new human respiratory isolate.

Pseudomonas strains produce rhamnolipid mixtures (RLs) that generally consist of one or two molecules of rhamnose linked to one or two molecules of 3-hydroxyalkanoic acid. This study evaluates carbon source effects (glycerol, glucose, myristic acid, and Brassica carinata oil) on the synthesis of monorhamnolipids (mono-RLs) versus dirhamnolipids (di-RLs) in a human isolate of Pseudomonas aeruginosa PAL05. Spectrophotometry, an emulsifying index (E24) test, and an orcinol assay confirmed the production of RLs by PAL05. Purified RLs were characterized by 1H NMR analysis. PAL05 primarily produces mono-RLs when provided carbon sources containing long chain fatty acids (FAs) (myristic acid and B. carinata oil) and di-RLs when provided glycerol or glucose. qRT-PCR analysis showed that delayed expression of rhlC occurred when B. carinata oil was used, but not glycerol, glucose, or myristic acid. Our data show that the carbon source influenced the transcriptional expression of the rhlC gene and, consequently, the predominance of mono-RLs or di-RLs in PAL05 cultures.

Raman spectroscopy differentiates between sensitive and resistant multiple myeloma cell lines.

Current methods for identifying neoplastic cells and discerning them from their normal counterparts are often nonspecific and biologically perturbing. Here, we show that single-cell micro-Raman spectroscopy can be used to discriminate between resistant and sensitive multiple myeloma cell lines based on their highly reproducible biomolecular spectral signatures. In order to demonstrate robustness of the proposed approach, we used two different cell lines of multiple myeloma, namely MM.1S and U266B1, and their counterparts MM.1R and U266/BTZ-R subtypes, resistant to dexamethasone and bortezomib, respectively. Then, micro-Raman spectroscopy provides an easily accurate and noninvasive method for cancer detection for both research and clinical environments. Characteristic peaks, mostly due to different DNA/RNA ratio, nucleic acids, lipids and protein concentrations, allow for discerning the sensitive and resistant subtypes. We also explored principal component analysis (PCA) for resistant cell identification and classification. Sensitive and resistant cells form distinct clusters that can be defined using just two principal components. The identification of drug-resistant cells by confocal micro-Raman spectroscopy is thus proposed as a clinical tool to assess the development of resistance to glucocorticoids and proteasome inhibitors in myeloma cells.

Sequestering ability to Cu2+ of a new bodipy-based dye and its behavior as in vitro fluorescent sensor.

A Bodipy (4,4-difluoro-4-bora-3a,4a-diaza-s-indacene) derivative has been conceived and synthesized starting from l-aspartic acid, as a selective turn-off sensor of Cu2+ ions. Its acid-base properties were determined to study the formation of metal/sensor complex species by titration of solutions each containing a different metal ion, such as Cu2+, Ca2+, Zn2+, Pb2+ and Hg2+ and different metal/sensor ratios. The speciation models allowed us to simulate the distribution of the metal/sensor complex species at the normal concentrations of the corresponding metals present in biological fluids. The distribution diagrams, obtained by varying the concentration of sensor 1, clearly indicate that sensor 1 responds selectively to Cu2+ at micromolar concentrations, even in the presence of other more abundant metal cations Ca2+. Finally, we analyzed the cellular uptake of sensor 1 on human erythrocytes and its ability to chelate Cu2+ in the cellular environment. Results indicate that it crosses the plasmatic membrane and colors the cells of a bright fluorescent red. Exposing the fluorescent cells to Cu2+ results in a complete cellular photobleaching of the red fluorescence, indicating that sensor 1 is able to detect metal changes in the cytosolic environment.

The role of glutamine in Pseudomonas mediterranea in biotechnological processes.

In this work, in order to study the effect of glutamine as co-feeder on growth kinetics, biomass and PHA production in Pseudomonas mediterranea, different co-metabolic strategies were employed. Unrelated (glycerol and glucose) and related (sodium octanoate) carbon sources both in presence and absence of glutamine have been tested. For each cultural condition, we (i) evaluated growth kinetics and measured the cell metabolic activity by MTT assay, (ii) monitored PHA production and (iii) estimated the expression of phaC1 and phaC2 genes through RT-PCR. Our results show that the use of glutamine as co-feeder in P. mediterranea led to an improvement of the specific growth rate and cell metabolic activity and enhanced the uptake of all the carbon sources assayed. Moreover, the use of glutamine reduced significantly the time required for PHA production and increased biopolymer yield, as consequence of an early activation of phaC1 and phaC2.

A micro-Raman spectroscopic investigation of leukemic U-937 cells in aged cultures.

Recently it has been shown that micro-Raman spectroscopy combined with multivariate analysis is able to discriminate among different types of tissues and tumoral cells by the detection of significant alterations and/or reorganizations of complex biological molecules, such as nucleic acids, lipids and proteins. Moreover, its use, being in principle a non-invasive technique, appears an interesting clinical tool for the evaluation of the therapeutical effects and of the disease progression. In this work we analyzed molecular changes in aged cultures of leukemia model U937 cells with respect to fresh cultures of the same cell line. In fact, structural variations of individual neoplastic cells on aging may lead to a heterogeneous data set, therefore falsifying confidence intervals, increasing error levels of analysis and consequently limiting the use of Raman spectroscopy analysis. We found that the observed morphological changes of U937 cells corresponded to well defined modifications of the Raman contributions in selected spectral regions, where markers of specific functional groups, useful to characterize the cell state, are present. A detailed subcellular analysis showed a change in cellular organization as a function of time, and correlated to a significant increase of apoptosis levels. Besides the aforementioned study, Raman spectra were used as input for principal component analysis (PCA) in order to detect and classify spectral changes among U937 cells.

Phage-AgNPs complex as SERS probe for U937 cell identification.

The early diagnosis of malignancy is the most critical factor for patient survival and the treatment of cancer. In particular, leukemic cells are highly heterogeneous, and there is a need to develop new rapid and accurate detection systems for early diagnosis and monitoring of minimal residual disease. This study reports the utilization of molecular networks consisting of entire bacteriophage structure, displaying specific peptides, directly assembled with silver nanoparticles as a new Surface Enhanced Raman Spectroscopy (SERS) probe for U937 cells identification in vitro. A 9-mer pVIII M13 phage display library is screened against U937 to identify peptides that selectively recognize these cells. Then, phage clone is assembled with silver nanoparticles and the resulting network is used to obtain a SERS signal on cell-type specific molecular targets. The proposed strategy could be a very sensitive tool for the design of biosensors for highly specific and selective identification of hematological cancer cells and for detection of minimal residual disease in a significant proportion of human blood malignancy.